
flass 1?:&37 
Book ,VVl 



Copyright N°_ 



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COPYRIGHT DEPOSIT. 



LABORATORY METHODS 



LABORATORY METHODS 



WITH SPECIAL REFERENCE TO THE NEEDS OF 



THE GENERAL PRACTITIONER 



BY 

B. G. R. WILLIAMS, M. D. 

MEMBER OP ILLINOIS STATE MEDICAL SOCIETY, AMERICAN MEDICAL ASSOCIATION, ETC. 

ASSISTED BY 

E. G. C. WILLIAMS, M. D. 

FORMERLY PATHOLOGIST OF NORTHERN MICHIGAN HOSPITAL FOR THE INSANE, 
TRAVERSE CITY, MICHIGAN 

WITH AN INTRODUCTION BY 

VICTOR C. VAUGHAN, M. D., LL. D. 

PROFESSOR OF HYGIENE AND PHYSIOLOGICAL CHEMISTRY AND DEAN OF THE DEPARTMENT 
OF MEDICINE AND SURGERY, UNIVERSITY OF MICHIGAN, ANN ARBOR, MICHIGAN 



SECOND EDITION 



ILLUSTRATED WITH FORTY-THREE ENGRAVINGS 



ST. LOUIS 

C. V. MOSBY COMPANY 
1913 






Copyright, 1913, by C. V. Mosby Company 



Press 0/ 

C. V. Mosby Company 

Si- Louis 



©CI.A350010 



TO 

THE GENERAL PRACTITIONER, 

WHO MUST BE A SPECIALIST IN ALL BRANCHES OF MEDICINE, 

THIS BOOK IS DEDICATED 

BY THE AUTHORS. 



PREFATORY NOTE 

A realization of the fact that the general practitioner is not 
usually prepared to make, on account of lack of extensive apparatus 
and other conveniences, elaborate chemical tests and examinations, 
has prompted the authors to prepare this book. "Laboratory 
Methods" is not of an encyclopedic form, nor is it a limited com- 
pend, and is especially designed for the general practitioner who 
desires to make, easily and inexpensively, examinations on which 
he may depend. 

The physician may have experienced some discouragement in 
attempts to conduct certain examinations, but he has probably 
been confused by the complexity of the large book and thwarted 
by the paucity of the compend. It is not presumed that the prac- 
titioner shall attempt every investigation, but this book will show 
that many of the comparatively simple cases that are usually sent 
to distant cities for expert examination may be made with more 
satisfactory results by the practitioner. 

It has been the aim of the authors to simplify methods both as 
to apparatus and technic. Essential factors have not, however, 
been omitted, but have been emphasized in such manner as to indi- 
cate their importance. Only the best tests are given, so that the 
reader will not be perplexed by being obliged to do any choosing 
for specific cases. Stress has been laid on safe diagnosis, and 
sources of error, as well as the value and limitation of tests, have 
been pointed out. 

B. G. R. W., M. D. 
E. G. C. W., M. D. 



PREFACE TO SECOND EDITION. 

Certain portions of this book have been reorganized, and there 
have been added descriptions of the Albumin Sputum Test for 
Tuberculosis, Bass and Watkins' Rapid Widal Method, Noguchi's 
Butyric Acid Test for Syphilis, and the Urobilinogen Test for 
Hepatic Function. 

Furthermore, an attempt has been made to meet some of the 
needs of the amateur analyst; and for this reason a few tests 
have been added in the form of an appendix, which includes a 
Bedside Method for the Estimation of the Urinary Acidity, a 
very valuable Indican Test, a simple Test for Indolacetic Acid, a 
consideration of the Bence-Jones Albumose Body, the Sulphosa- 
licylic Acid Test for Urinary Albumin, and the Hermann-Perutz 
Serum Test for Syphilis. 

B. G. R. W., M.D. 
E. G. C. W., M. D. 
April, 1913. 



CONTENTS 



CHAPTER I. 

PAGE 

General Considerations 13 

CHAPTER II. 
The Sputum 27 

CHAPTER III. 
Searching for Germs 3/ 

CHAPTER IV. 
Vascular Dramas 53 

CHAPTER V. 
Chemistry and Biology of the Gastric Juice 71 

CHAPTER VI. 
Essence of Tissue Diagnosis „ .... 78 

CHAPTER VII. 
Detection of the Common Poisons 85 

CHAPTER VIII. 
Exudates in Brief 94 

CHAPTER IX. 
Diazo Versus Widal 100 

CHAPTER X. 
The Urine in Disease 107 

_ CHAPTER XL 
Milk and Its Home Modifications . . . . . „ 125 

CHAPTER XII. 

Some Simple Water Analyses 138 

7 



O CONTENTS 

CHAPTER XIII. 

PAGE 

Eveby-Day Stool Tests , . „ . 148 

CHAPTER XIV. 
Technic of the Pkivate Post-Mobtem 156 

CHAPTER XV. 
To Find the Treponema in Six Minutes . .171 

CHAPTER XVI. 
Laboratory Prophylaxis 177 

CHAPTER XVII. 
Indications foe Labobatoby Aids 182 

CHAPTER XVIII. 
Genebal Infobmation « 186 



ILLUSTRATIONS. 



FIG. PAGE 

1 Giant alcohol burner 17 

2 Kerosene mantle burner 17 

3 Portable microscope 19 

4 Physician's laboratory 22 

5 Apparatus for sputum analysis 28 

6 Elastic tissue and sputum findings 30 

7 Apparatus for bacteriological examinations 36 

8 A streak culture 38 

9 Correct method of inoculating a tube 39 

10 Surface colonies of diphtheria bacillus 44 

11 Some of the more common forms of the diphtheria bacillus . . .45 

12 Typical fir tree tetanus stab in gelatin 46 

13 Apparatus for blood examinations 52 

14 Development of the blood cells ' 54 

15 Scheme for rapid counting of red cells 61 

16 Spreading 63 

17 Artifacts in blood films 64 

18 Authors' slide forceps 66 

19 Apparatus for stomach analysis 70 

20 Method of removing contents from stomach 73 

21 Microscopic elements of major and minor import in stomach analysis 74 

22 Substitutes for the perfected microtome 79 

23 Apparatus for detection of common poisons 86 

24 Improvised Marsh apparatus 89 

25 Puncture of pleura 95 

26 Diagram showing site of lumbar puncture 96 

27 Apparatus for urinalysis 106 

28 Modification of Boston's pipette test for albumin Ill 

29 Artifacts . . . . .' 118 

30 Some typical epithelial cells from the urinary passages . . . .119 

31 Comparison of the more usual forms of the common crystals met in 

urinary sediments 120 

32 Tube for fat estimations 135 

9 



10 ILLUSTRATIONS. 

FIG. PAGE 

33 Tapeworms compared with vegetable fibers 151 

34 Ova of the most common intestinal worms compared with a red blood 

cell 152 

35 Scalp incision 160 

36 Removing the skull cap 162 

37 Initial incision 164 

38 1 Examination of the heart 165 

39 Apparatus for finding treponema pallidum ........ 170 

40 Making the suspension 172 

41 Spreading the mixture 173 

42 Treponema pallidum 175 

43 Five-inch test tube, with various amounts of liquid . 186 



INTRODUCTION. 

BY VICTOR C. VAUGHAN, M. D. 

It gives me great pleasure to write a short note of approval of 
this book. It has been said that the general practitioner is passing 
away, but this statement is certainly negatived by this book. There 
is nothing more hopeful in the practice of medicine today than the 
thorough way in which many general practitioners are doing their 
work. There are many small cities, and even villages, in which 
there are general practitioners who have equipped themselves with 
most effective laboratories. This volume shows that the working 
laboratory in which the best work may be done can be established 
at a small cost. It requires only a good man to conduct it. It 
would be regrettable were it true that the country doctor has ceased 
to do scientific work. Jenner was a village doctor when he tested 
and demonstrated the efficiency of vaccination for smallpox. Pas- 
teur had shown himself a great scientist before he ever saw Paris. 
Koch was a stabsarzt, remote from any great medical center, when 
he devised solid culture media for the growth of bacteria,* and 
opened up a method of scientific investigation which has given such 
brilliant results. Sims was a practitioner in the then village of 
Montgomery, Alabama, when he worked out the technic of the suc- 
cessful operation for vesico-vaginal fistula. Long was a rural doc- 
tor in Georgia when he first removed a tumor under general anes- 
thesia. Pollender was a country doctor when he first studied the 
blood of animals sick with anthrax, and demonstrated rod-like 
organisms in the same. Beaumont was an army surgeon, stationed 
at an isolated post on the Island of Mackinac, in the then territory 
of Michigan, when he made his now classical experiments upon 
Alexis St. Martin. Indeed, if we take away from medicine the con- 
tributions to that science made by physicians far removed from 
great commercial centers, we rob it of half its glory and its honor. 

This little volume shows how the general practitioner can, at a 
very small cost, equip a laboratory in which he can do most excel- 

11 



12 INTRODUCTION. 

lent work. It demonstrates that costly apparatus and marble 
rooms are not necessary for the prosecution of scientific medicine. 

It gives me great pleasure, after a careful reading of the proof, 
to commend most highly this volume. 

Ann Arbor, Michigan. 



LABORATORY METHODS. 



CHAPTER I. 

GENERAL CONSIDERATIONS. 

McDowell, who performed the first ovariotomy, had not at his 
elbow the nickel-plated sterilizer and the gowned assistant, but to 
his ears came the mutterings of a mob which had sworn to take 
his life if he failed in the operation. Sims worked and enjoyed 
his labors under circumstances that would have deterred many 
physicians, and Beaumont had not the advantage of even an occa- 
sional visit to the city clinics. 

Medical analyses, meaning those procedures where chemistry and 
microscopy are used to aid in making correct diagnoses, have gained 
an important position — too important to be ignored or turned 
mutely over to the expert. 

LABORATORY EQUIPMENT. 

When purchasing laboratory equipment, it is advisable to pro- 
ceed carefully. Too many laboratories, especially those which fail 
to give satisfactory results, are fitted too hurriedly. While it is 
not suggested to pay high prices for a popular trademark, some 
very worthless apparatus is being sold. Stains, as well as solu- 
tions for quantitative work, should be purchased in liquid form 
ready for use, and a number of reputable firms prepare good 
reagents. 

In this book, where possible, the English system of weights and 
measures has been used. 

The authors have listed several departments in order that the 
various needs of physicians may be met, as it is much better to 
become expert in blood analyses alone than to conduct incompletely 
several lines of work. 

13 



14 LABORATORY METHODS. 

Microscope. — The microscope, once a luxury, is now imperative 
for a safe diagnosis. It may be purchased for less than the more 
elegant examining chairs, and is, to say the least, just as neces- 
sary. A good instrument may be had for $80, this price including 
the third or oil immersion objective — not a necessity, but certainly 
a convenience. For ordinary work a movable stage is unnecessary, 
though many men who have not been forced to work without it 
would not agree with this conclusion. It is well to remember that 
Americans make good lenses. 

Glassware. — A few cents will buy a stock cf glass tubing and 
stirring rods, and gray filter paper is cheap. A glass funnel and 
one or two graduates are very convenient. 

Centrifuge. — A hand centrifuge, with extra milk tube, may be 
purchased at a low figure. Sedimentation glasses are possible, 
though not ideal, substitutes, and when collecting urinary sedi- 
ments a little thymol should be added to prevent fermentation. 
Not only does the centrifuge save time, but serves to ' ' bring 
down" elements which would otherwise remain suspended in the 
sedimentation glass. 

Slides. — Glass slides and covers are obtainable at any physicians' 
supply house, and cost very little. For the finer work these slides 
should not be too thick. On the other hand, a piece of window 
glass 2x2 inches serves well for urine examinations. Slides which 
have been used should not be cast aside as worthless. If per- 
manent preparations have not been made, a little alkali solution 
will remove smears of bacteria or blood. If these have been fixed 
or stained, the solution may be made stronger. Balsam may be 
removed by xylol. A dip in grain alcohol may precede washing 
in water. 

Accessories. — The following are some of the laboratory acces- 
sories, with prices : 

File for glass work 05 

Platinum wire 30 

Evaporating dish 15 

Eing stand equipped $1.00 

The selection of rings for the ring stand should include a special 
clamp to serve as a buret support, 

Cleaning Glassware. — Money and time will be saved by cleanli- 
ness. For glassware, hot suds followed by hot water are best. 



GENERAL CONSIDERATIONS. 15 

Polish with a soft, dry cloth. Strong mineral acids will remove 
organic matter. 

Stains. — Wright's blood stain, carbol-gentian violet, fuchsin, and 
methylene blue are necessary for the blood and bacteriological 
work. For tissues, carbol thionin gives a beautiful effect. Hema- 
lum or alum carmine may be substituted, and then the prepara- 
tion stained with eosin if a beautiful permanent mount is desired. 
Alcohol, carbol-xylol, and balsam are necessary only in case the 
preparation is to be filed. Balsam should not be bought in bottles, 
but in tubes, in order to prevent waste and decoloration as well as 
contamination, etc. A paper-filtered balsam is most transparent. 
A spatula or section lifter may prevent the ruining of many thin 
sections. It is well to remember that in case the stain should be 
overturned and spilled, recourse may be had to certain anilin inks. 
For this purpose eosin, gentian violet, fuchsin, and certain other 
stains may be obtained in every hamlet, and extraneous matter 
removed by filtration. Good liquid stains should, however, be used 
when possible. Certain fabric dyes have been used to demonstrate 
the gonococcus. India ink and its American substitutes do not 
stain microorganisms, but have been recommended as a substitute 
for the high-priced dark field attachments in searching for the 
treponema pallidum. 

Substitutes. — Common blotters may be used to take up excess 
liquids. A brandy bottle makes a good flask unless contents are 
to be boiled, a procedure rarely or never necessary in clinical 
analyses. Whisky goblets may be used as beakers when titrating 
or as wash dishes, while saucers serve as watch glasses. Small 
new tin ointment boxes, such as are used in dispensing, may be 
substituted unless acids are used, the bright background forming 
an excellent contrast to the floating sections. Two-ounce bottles — 
round or square, to prevent overturning — serve well for liquid 
reagents. Small salt-mouth bottles or granule bottles are best for 
solids. Raised glass labels are unnecessary, but glass stoppers, 
especially for the stronger acids and alkalies, are imperative. It 
is advisable to keep only' a small quantity of the reagent in the 
bottle which is to be used in order to guard against evaporation, 
contamination, or spilling. Stock reagents should be kept in a 
dry basement during the summer months, and never exposed to 
the light. 

Indicators. — A dilute solution of phenolphthalein is the cheap- 



16 LABORATORY METHODS. 

est, as well as the best, chemical indicator. When the solution is 
acid or neutral, it is colorless. Alkalinity results in a red colora- 
tion. Litmus paper is too crude for medical work. 

Cover Glasses. — Round cover glasses are well adapted for smears, 
but the squares are more convenient for tissue sections. They 
may be cleaned by immersion in a little alcohol, rinsing in hot 
water, and polishing with a dry, soft cloth. A ' ' linty rag ' ' should 
be avoided, as vegetable fibers do not add to the beauty of the 
miscroscopic field. It does not follow, however, that in the event 
such contamination occurs the physician should confuse these with 
elastic fibers or urinary casts. 

Pipettes. — Medicine droppers serve well as pipettes, both for 
staining and for transferring liquids. One delicately graduated 
pipette is necessary for stomach work. 

Apparatus Cabinet. — An old bookcase will serve well to keep 
dust off apparatus, as a general house cleaning preliminary to a 
test should not be necessary. The inside of this case should be 
painted white and the outside black. 

Laboratory Nostrums. — The "general stain-all," the "glass 
cleaners," and many other preparations offered to the indifferent 
practitioner may be aptly termed "laboratory nostrums." The 
secret "blend" and the advanced price seem to be the only dis- 
tinctions from those efficient formulas well known to all scientific 
students. The urinary test tablets should b>e avoided. 

Suggestions. — In each chapter helpful suggestions have been 
made in regard to good, but not expensive, apparatus. The diffi- 
culties in technic most liable to be met by the practitioner are 
pointed out, and methods suggested as to how these may be over- 
come. 

Stock analytic outfits selected by supply houses should not be 
purchased. The urinary analysis hand case will rarely, if ever, 
be taken from the physician's office, and is not adapted for prac- 
tical work. Such selection should be made as will meet the re- 
quirements of the individual physician, reagents should be obtained 
in fresh condition, and both time and money will be saved. 

Appropriate substitutes for the unavailable articles of equipment 
will constantly suggest themselves to the resourceful man, and the 
physician who, in an emergency, finds that he may inoculate a 
tube by means of a hat pin instead of a platinum point brings as 
much honor to the disciples of Esculapius as does he who ampu- 



GENERAL CONSIDERATIONS. 



17 



tates with butcher knife and meat saw the gangrenous limb of the 
frontiersman. As in other branches of medicine, intellect first — 
equipment later. 





Fig. 1. — Giant alcohol burner. Its flame 
is four times as intense as that of 
the ordinary Bunsen gas burner. 



Substitutes for Gas and Running Water. 

Gas. — The authors have demonstrated that gas is not only un- 
necessary, but sometimes undesirable, in laboratory work. In the 
large laboratory it is convenient, 
and, when satisfactory, should be 
used. The country physician has 
recourse to alcohol and gasoline. 
The small spirit lamp is inexpen- 
sive, may be taken to the bedside, 
uses only a small quantity of alco- 
hol, and does not place him at the 
mercy of some gas plant engineer. 
There will be no "water in the 
pipes," no "cut outs," and no 
"meters." Various modifications 
of the alcohol lamp are marketed. 
A portable burner 
with a wall tank can 

be had (Fig. 1), the reservoir holding about a quart 
of denatured alcohol, from which a flame four times 
as intense as that of the ordinary Bunsen burner 
may be obtained. 

For continuous heating, a single-burner gasoline 
stove is ideal. A kerosene burner fails as a sub- 
stitute, as its flame is usually inaccessible. A uro- 
tropin tablet, burned in the air, will heat to boiling 
the contents of three test tubes successively ap- 
plied. This tablet should be placed on a glass or 
metallic plate, or in a medicine spoon. Poor grades 
of this drug have, however, a tendency to explode; 
throwing burning particles in all directions. 

Gas is not necessary for incubators (page 21), 
and neither is it required for illuminating pur- 
poses. By properly manipulating the mirror, con- 
Fi mantle"£vn?r ne denser, an d diaphragm, smears and sections may be 



18 LABORATORY METHODS. 

examined by the ordinary kerosene lamp. The authors have tested 
with the most gratifying* results the various mantle kerosene 
burners, one of which is shown in Fig. 2, and do not hesitate to 
recommend them to all microscopists as the best, and yet the most 
economical, of lights. 

Running Water. — A city water supply may be aptly termed a 
laboratory luxury, and, when obtainable, is not to be despised. A 
siphon system, operated from a large bottle on a shelf, with a rub- 
ber tube and a pinch cock, will, however, answer very well. In 
staining, wash glasses (whisky glasses containing water) will save 
many steps. A large bowl of clean water serves well for washing 
sections and smears, as anilin stains, when much diluted, are prac- 
tically inert. Blood pipettes may be cleaned without the aid of 
running water (see page 62). Centrifugalization need not depend 
on water pressure. 

Essentials of Practical Microscopic Technic. 

Mirror. — The plane mirror is the one usually employed, espe- 
cially when examining stained preparations. The concave mirror 
is preferred when using artificial light or when examining tissue 
sections. In the latter case it is best to swing aside the condenser 
and narrow the aperture of the iris diaphragm. 

Illumination. — Direct sunlight should never strike the mirror. 
The best angle for the reflection of light is as it comes from a 
white cloud (Novy). A white window shade will often be a great 
aid, especially in a south room. 

Condenser. — Fig. 3 illustrates the portable microscope. The 
stage has been tilted in such a manner that the condenser and its 
control B are seen to best advantage. The use of the condenser 
usually comes with experience, but it should always be adjusted 
when beginning work. The man who "plays" with his condenser 
will soon become expert in its use. A correct focus of the con- 
denser is as important as the objective focus. Neither increases 
the magnification, but both serve to render distinct the smear or 
section. 

Iris Diaphragm. — This is rarely, if ever, opened wide. Correc- 
tion of illumination is very often necessary in seeming indications 
for a widened aperture. With a slightly constricted diaphragm a 
stained object is rendered more distinct, and, when examining un~ 



GENERAL CONSIDERATIONS. 



19 




Fig. 3. — Portable microscope. A, to adjust draw tube; B, to control condenser: C. dia- 
phragm; D, coarse adjustment; E, fine adjustment. 



stained preparations^ further narrowing is necessary, as that field 
is bathed in a dim twilight, the best possible illumination for such 
examinations. 

Ocular. — This regulates, to some extent, the magnification. The 



20 LABORATORY METHODS. 

various oculars are numbered differently according to the make of 
the instrument. It is of advantage in diagnostic work to change 
oculars frequently. 

Objective. — The use of the objective is usually so well under- 
stood that its description is omitted. The objective perfects the 
image, the latter lying just below the ocular. 

Focusing. — This does not mean merely a lowering or raising of 
objective, but also an adjustment of the condenser. It is a good 
plan to make the final adjustment of the condenser after the objec- 
tive is in proper position in order to obtain the most perfect image. 
It is usually advantageous to begin the study of a preparation with 
the lowest power and then proceed to higher magnifications. A 
good procedure for "safe" focusing is offered by Stitt: "It 
should be an invariable rule for the worker to bring his objective 
practically into contact with the upper surface of the cover glass, 
using the coarse adjustment to slowdy elevate it into focus, and 
then maintain this focus with the micrometer screw." 

Cleaning Lenses. — Expensive lens paper is unnecessary. An un- 
starched, but clean, linen handkerchief is much better. Oils are 
best removed with the aid of a similar cloth previously moistened 
in a little alcohol ; if, however, it is applied to the tube, the alcohol 
quickly destroys the lacquer. "Warm water and a soft cloth re- 
move agar-agar and gelatin. Balsam is not so easily conquered, 
and must be avoided. Carbol-xylol will dissolve it, but its fre- 
quent use dissolves the cement which secures the lenses and may 
eventually loosen them. 

Cover Glasses. — These should be used invariably, especially when 
working with the higher powers, as they render the object much 
more distinct. Prior to their use a drop of water or balsam must 
be placed on the preparation. Thick cover glasses interfere with 
high-power focusing and must not be used. Round cover glasses 
are used for smears of pus ; for tissue section squares are preferable. 

Microscopic Hysteria (Microscopic Cephalalgia) .—This is not 
usually an idiosyncrasy, but results from one or more well-known 
causes. It may be due to an uncorrected error of refraction, to 
improper illumination, malposition of condenser, or an open 
diaphragm. A severe headache may arise from an accommodation 
squint. If one eye is closed, it soon becomes fatigued because the 
working eye must accommodate. A dark glass worn over the un- 
employed eye will often relieve this condition. Both eyes should 



GENERAL CONSIDERATIONS. 21 

be open. It is a good plan to alternate the use of one eye with 
the other. 

Laboratory Tables. — These should be painted black, but walls 
should be of a light tint. 

Miscellaneous Rules. — The following rules should be carefully 
observed. 

1. Use microscope in a vertical position, and slide clamps will 
not be necessary. 

2. Use a chamois skin to polish lacquer. 

3. Keep fingers off lacquer, and carry the microscope by its 
handle. 

4. Do not lay a slide on the stage until its under surface is 
known to be dry. 

5. A microscope, when not in use, must be kept from dust by 
placing it either in its case or under a bell jar. 

A Bacteriological Laboratory for Five Dollars. 

Five dollars or less will buy everything necessary for a phy- 
sician's bacteriological laboratory — microscope not included. The 
bugbear in considering such a laboratory has been the incubator, 
and this matter has usually been presented to the general prac- 
titioner in a discouraging manner. The fact is, however, that all 
that is necessary is some method of maintaining cultures at a con- 
stant temperature of 98.6° F. — i. e., at blood heat. 

During the hot summer months no such contrivance is necessary, 
and for other times of the year many substitutes for the $50 gas 
incubator may be made. For example, a chicken incubator will 
serve the same purpose, a kerosene "germ warmer" that will an- 
swer is on the market, and a high candle power electric bulb may 
be immersed in a pan of water and the heat regulated by adding 
or removing water. A certain poultryman was able to hatch 
chicks in a bee hive from the heat given off by these insects, and, 
although the establishment of an apiary in a clinical room is not 
recommended, the method is not without a lesson. Perfect cul- 
tures of the diphtheria bacillus have been obtained in an office 
slightly overheated by an ordinary round stove, and an instance 
is known where a man carried a living culture of the typhoid 
bacillus safely through a blizzard by means of a special pocket in 
his underwear. A considerable fall in temperature, though inhib- 



22 



LABORATORY METHODS. 




-« Bf 



A hT 



^2 _ 



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ft£ . 

w £ fl 



O) ft 

c ° 



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o .. k 
» n ^ 



to oa 

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GENERAL CONSIDERATIONS. 23 

iting the growth of many microorganisms, does not necessarily 
result in their death. These facts account for the possibility of 
mailing diphtheritic material to far distant laboratories, the reten- 
tion of the virulence of "spoiled" ice cream, etc. 

A tireless cooker will serve to keep plate cultures at a growing 
temperature, and for tubes the vacuum bottle has been recom- 
mended, which is not only convenient, but has the advantage of 
being well adapted for refrigerative purposes. 

The hot air sterilizer may be easily dispensed with, as there is 
no better sterilizer for test tubes, metal ware, flasks, and cover 
glasses than a small gasoline oven, which may be heated with a 
gasoline or alcohol burner. 

Culture media should be obtained from a reputable firm in 
assorted lots of a dozen, which will save considerable expense and 
trouble that would be incurred in an attempt to prepare them. 
After using these media the cultures may be killed and the tubes 
cleaned for chemical analyses. 

The following estimate of prices of accessories should be con- 
sidered by the practitioner : 

Vacuum bottle $1.95 

Gasoline oven 1.25 

Alcohol stove 1.75 

Six tubes of culture media 30 

Platinum loop 30 

$5.55 

Arrangement for a Physician's Laboratory. 

The idea that the clinical laboratory must be a separate institu- 
tion is erroneous. Its location need not be seclusive, nor need it 
be even separated from the consultation room. One side of the 
ordinary examining room may be devoted to laboratory purposes, 
and the light side should, of course, be chosen. An ideal arrange- 
ment is shown in Fig. 4. 

A shelf table, without .legs, should be made to run the entire length 
of the laboratory space. Cypress is the best w T ood for its construc- 
tion, and it should measure about 20 inches in width. It should 
be at a height convenient for the "standing" analysis, and a 
high stool, when desirable, may be used for the microscopic examina- 
tions. 



24 LABORATORY METHODS. 

At one end of this shelf table should be located the apparatus 
cabinet, and the sink is built into the other end. All apparatus 
should be kept free from dust in the cabinet, at least a portion of 
which should be covered with glass. 

The shelf table should be securely fastened to the wall in order 
that there be no legs to trip the foot or thwart the broom, and a 
small quantity of black paint will add the finishing touches. Such 
a laboratory table will require little or no room, as examining 
chairs, bandage or dressing tables, etc., may be placed under it 
when these are not in use. 

In case sunlight is direct and blinding, white shades may be 
drawn over the windows. 

The shelf can be easily kept clean with a chamois, slightly damp- 
ened, and formalin added to the water not only inhibits or kills 
pathogenic microorganisms, but will destroy odors of pus, urine, 
etc. 

Offensive specimens of excretions or bits of diseased tissue should 
be kept from the view of patients. 

During a urinalysis the samples should be kept in amber or blue- 
colored glass bottles or jars. A "stinking" sample of urine is 
rarely excusable. Except in some forms of cystitis or gynecologic 
conditions, a freshly voided urine never smells. If, however, an 
analysis of a bad-smelling specimen is necessary, it should be de- 
ferred until an hour when visitors are most unlikely to enter. 
After such analysis there should be a thorough aeration of the 
room, for which purpose the following formula is a most excellent 
deodorizer : 



E Iodoform % j 

Oil of spearmint ad saturated solution 

Sig. : Nebulize, vaporize, or otherwise distribute throughout the 
room. 



The physician should remember that most reagents are freezable, 
with the following notable exceptions : carbol-xylol ; alcohol and 
alcoholic stains and solutions; ether; solutions containing glycerin 
— for example, Haines' solution. 

Laboratory reference books should be kept with the laboratory 
equipment, ready for convenient reference. This book was not 
written for the bookcase, but was intended to lie on the table 



GENERAL CONSIDERATIONS. 25 

within easy reach of the physician. On the laboratory walls may 
be hung charts of bacteria, solubility tables, etc. 

.Most solutions and stains should be kept only in small quantities 
on account of evaporation, precipitation, etc. Haines' solution 
should be prepared fresh in small amounts every few months. 

Further laboratory suggestions have been made in succeeding 
chapters. For example, directions for the care of blood pipettes 
occur in Vascular Dramas (page 62), principles of asepsis and 
antisepsis occur in Searching for Germs (page 42), etc. 

LABORATORY EXPERTS. 

Before giving a descripticn of the various teste it seems advisable 
to sound a word of warning in regard to a subject about which 
there seems to be much confusion — laboratory experts. Modern 
medicine, especially its laboratory branches, is built on a scientific 
foundation; in fact, its progress is a history of the progress of 
prophylaxis and diagnostics, each physician having, to some extent, 
his own ideas regarding therapeutics. There was a time when a 
certain mysticism was associated with sending a specimen to some 
prominent chemist for a test, but now medical students are trained 
to perform and understand these tests. A physician should per- 
form all the tests which present knowledge and equipment place 
within his reach; but what shall he do when he is not prepared 
to make some important investigation ? 

Practitioners are cautioned against some laboratories. To state 
a diagnosis to a patient means to pass judgment, and is a serious 
matter. In case the physician has not the time or is not prepared 
for a certain investigation, he should exercise care as to whom he 
calls into consultation. In the first place, he should studiously 
avoid the ordinary chemist or the chemical supply house, and 
should seek the medical specialist. The institution which asks no 
remuneration for its services should be avoided, although this may 
not be advisable when dealing with patients unable to pay the 
necessary fees. A curt' "positive" or "negative" from some 
student, or from a favorite of unknown ability, not only means 
nothing to the conscientious therapeutist, but such report is usually 
so misleading as to be dangerous. Indeed, if as much care were 
taken in the proper selection of consultants in diagnostics as in 
the field of surgery, or in the department* «f ophthalmology and 



26 LABORATORY METHODS. 

otology, our medical successes would more than compensate for 
such efforts. 

It may be well before leaving this subject to caution physicians 
against the sending of specimens to sanitariums or other institu- 
tions whose ultimate aim may be to secure for themselves the pa- 
tients from whom the specimens were taken. The same caution 
will apply to reports from tent colonies concerning sputum exami- 
nations or to reports from certain mineral springs in regard to 
urinalyses, as these reports are liable to be biased and their recom- 
mendations be accordingly unsafe. 

The physician should personally do as much as possible of his 
analytical work, and should spare neither time, money, nor study 
to become a safe diagnostician. The practical information pre- 
sented in this book will aid him in his endeavors, and, in case 
expert help becomes necessary, the best help obtainable should be 
secured. Patients will appreciate intelligent treatment and will 
be willing to pay accordingly. 



CHAPTER II. 

THE SPUTUM. 
Apparatus. — Does not include that used in the albumin test. 



1. Glass plate. 

2. Teasing needles (sharp hat pins 

make excellent substitutes). 

3. Platinum loop. 

4. Black laboratory table or card 

board. 

5. Slides. 

6. Round cover glasses. 



7. Carbol-gentian violet. 

8. Dilute nitric acid. 

9. Very dilute acid fuchsin. 

10. Ordinary blotting papers. 

11. Several whisky glasses. 

12. Cover glass forceps. 

13. Flame. 

14. Microscope and accessories. 



Scope of Work. — The arrangement of apparatus is shown in 
Fig. 5. Those tests will be described which are most commonly 
needed by the practitioner, and less frequently applied procedures, 
including those usually left to experts, are merely listed. 

Obtaining the Sputum. — Give the patient 15 grains of potassium 
iodid at bedtime, and, if possible, repeat this dose at about 2 
o'clock the following morning. This drug is preferably given in 
milk. On rising in the morning the patient should wash out his 
mouth with a little soda water, being careful not to hawk up at 
the time any sputum. The sputum is then collected in a wide- 
mouth vaselin bottle. Saliva is not wanted. Sputum should be 
coughed up or raised by hawking. It is manifestly a waste of 
time to examine oral, nasal, and pharyngeal secretions for evidences 
of pulmonary involvement. 

Quantity. — This is so variable in the several stages of lung dis- 
eases as to be of little diagnostic import. A very small quantity 
of true sputum is the rule in incipient tuberculosis. 

Consistence. — Here, again, are variations. When the sputum is 
very thin and watery, showing a tendency to froth, there is a 
probability of pulmonary edema. Pus is easily divided into drops 
with the aid of a pipette, but mucus has a tendency to cling, not 



References. — Sahli : Diagnostic Methods; Von Jaksch: Diagnostics; Boston: Clinical 
Diagnosis; Wood: Chemical and Microscopical Diagnosis. 

27 



28 



LABORATORY METHODS. 




THE SPUTUM. 29 

only to other matter (adhesion), but is separated into drops with 
difficulty (cohesion). 
Color. 

Color. Cause. Significance. 

Translucent and slimy. . Mucus Variable. 

Black Soot Normal in soma occupa- 
tions. 
Rusty, prune juice, or 

yellow Changed blood Usually pathological- 

Green Hemorrhage or icterus. . Pathological. 

Gray Pus Pathological. 

Red Blood Pathological. 

Small clumps of a bright-green color may often be observed in 
influenza. 

Odor. — Sputum is odorless, except when it becomes contami- 
nated with putrefying microorganisms, which may be observed 
in pulmonary gangrene or bronchiectasis. Sometimes a sputum, 
after standing for a few hours, unless kept on ice, becomes 
soured. The authors have seen a drop of secretion from a fetid 
rhinitis contaminate an odorless sputum to such an extent as 
to render examination almost impossible. 

Albumin Sputum Test. — Soluble albumin in the sputum is 
identified in much the same manner as in urine, but because of 
the presence of mucus, specimens of sputum are filtered with dif- 
ficulty, so that it is necessary to modify the technic somewhat. 
In a large test tube, five drams of physiologic salt solution, five 
c.c. of sputum and five drops of acetic acid, are well shaken for 
five minutes. It is easy to commit to memory this technic when 
the quantities are thus remembered in fives, though the measure- 
ments do not need to be accurate. Shake well, filter and test the 
filtrate for albumin by any of the tests which may be applied for 
serum albumin in the urine. In chronic cases where uncompen- 
sated valvular lesions and nephritis may be ruled out, the posi- 
tive reaction invariably means tuberculous involvement of the 
lungs as albumin is not met in chronic bronchitis. It is usually 
present in acute diseases of the lungs and bronchi, and is of 
practically no differential value except in protracted conditions. 

Searching for Elastic Tissue. — Pour all the sputum on the glass 
plate and set this plate on a black surface for examination. A 



30 



LABORATORY METHODS. 



laboratory table painted black will answer the purpose. Next 
tease out the solid portions of the specimen with needles. Bits of 
elastic tissue — i. e., lung tissue — appear usually as yellowish gran- 
ules. Very rarely they may be tinged with red blood or may be 
hardened (calcification). These bits of tissue should be selected 




Fig. 6. — Elastic tissue and other sputum findings with which it may be confused. A, 
elastic fibers, with acute branchings and ends resembling fish-hooks, seen only when 
diaphragm is narrowed, and in a moderate light they show a double contour ; B, 
fibrinous bror>chial cast; C, molds; D, mucous spiral; E, cotton fibers, extraneous, 
and usually come from the towel used in wiping laboratory glassware; F, linen fibers. 



out, and examined for the typical yellow fibers. Low power should 
be used, and, |f the field is well illuminated, the diaphragm should 
be somewhat narrowed. Food particles, actinomycotic granules, 
tonsil plugs, etc., may be mistaken for these particles of lung tissue. 
Under low power the elastic fibers refract light in such a manner 
as to make them appear double, which can not be properly shown 
in a drawing. In Fig. 6 elastic fibers are shown, and their ap- 
pearance is so characteristic that confusion with the other ele- 
ments in this figure is rarely excusable. Their broken ends tend 



THE SPUTUM. 31 

to curl into a form not unlike that of a fish-hook, and branching 
at acute angles may also be noted. When once recognized, they 
do not usually cause confusion in future examinations. 

Significance of Elastic Tissue. — Its presence in sputum means, 
invariably, the coughing up of diseased lung tissue, and signifies 
usually the presence of Koch's bacillus. Elastic tissue may some- 
times be found in nontuberculous abscess, rarely in pulmonary 
gangrene, and never in normal sputum. 

Preparing the Spreads. — Pick out suspicious particles with a 
platinum loop which has been previously sterilized in the flame 
and then cooled. Spread these well on clean round cover glasses 
and allow them to dry. Do not neglect to heat the loop to redness 
before laying it aside. When thoroughly dried, pass these cover 
glasses, with the specimen side up, rapidly through the flame sev- 
eral times, taking care not to burn the preparation. A brownish 
coloration indicates that the preparation has been ruined. The 
cover glass should, however, feel hot to the finger, and this heat 
fixation not only glues the preparation to the glass, but causes the 
stain to hold. 

Staining the Preparation. — The following modification of the 
Ziehl-Neelsen method and its congeners has given the authors the 
best results : 1 

Seize the cover slip with forceps and lock. Hold specimen side 
up, and cover it with a few drops of carbol-gentian violet. Hold 
above the flame in such a manner that it steams, but does not boil. 
This height is soon learned, and varies with the intensity of the 
flame. Replace with a medicine dropper any of the stain lost by 
evaporation. After steaming for three minutes, the excess stain 
is poured off and the preparation is ready to go through the 
washes. The latter are contained, most advantageously, in small 
tumblers, and the actual process is that of paddling gently each 
different liquid with the preparation. Each specimen should go 
through the following washes : 

1. Water, one minute. 

2. Dilute nitric acid until only a bluish tint remains. 

3. Ethyl alcohol, 50-percent, two seconds. 

4. Water immediately (not that used in 1), two minutes. 



1 This same technic is applicable where the physician prefers to stain with carbol- 
fuchsin and counterstain with methylene blue or bismarck brown. It is occasionally ad- 
visable to apply both methods to the same specimen in case any doubt exists. 



32 LABORATORY METHODS. 

5. Acid fuchsin solution, one-half minute. 

6. Water, two minutes. 

Dry the preparation between two ordinary blotters and examine, 
mounted either in water or balsam. The tubercle bacilli should 
appear as small, slender violet rods on a pink background. Oil 
immersion objective should be used in the examination. 

To Prepare Dilute Nitric Acid. — Add four drops of concen- 
trated nitric acid to each half ounce of water. 

Oarbol- Gentian Violet. — Use the liquid stain as prepared by 
some responsible company. 

To Prepare Acid Fuchsin Solution. — Use 1 part of the alcoholic 
solution to 20 parts of water. Filter if necessary. Dilute picric 
acid makes even a more beautiful counterstain, but does not stain 
properly other microorganisms. It should be used only by one 
who has learned to recognize the tubercle bacillus by its mor- 
phology as quickly as by its staining characteristics and who has 
thoroughly mastered the technic. 

The Findings. — It is not sufficient that Koch's bacillus is absent 
or present. Is there a secondary infection, or is any purulent in- 
fection which may be present the primary condition ? What germ 
is causing the symptoms? What is its degree of virulence ? What 
stage of tuberculosis is present? Is there caseation or lapidifica- 
tion? Types of white blood cells are of some import, and many 
polymorphonuclears are more apt to indicate secondary infection 
than a few lymphocytes. Records of over one thousand analyses 
made by the authors indicate that correct diagnostic and prog- 
nostic conclusions were often obtained from corpuscular elements 
alone. The following table illustrates some of these findings : 

Phthisis. Purulent infections. 

Elastic tissue None. 

Lapidification (lung stones) None. 

Bacillus tuberculosis Nonacid fast cocci or bacilli, among 

which certain types usually predomi- 
nate. 

Caseous masses None. 

Blood (red cells) None. 

Lymphocytes Pus cells — i. e., disintegrating poly- 
morphonuclear leukocytes. 

Appearance of the Tubercle Bacilli. — These tend to occur in 
clumps. Each measures in length about one-half the diameter of 



THE SPUTUM. 33 

a red blood cell, and often presents a beaded appearance or shows 
a slight curving of the ends. While branched forms have been 
observed, they rarely branch. In very virulent strains they may 
appear to be shortened even as coccus-like bodies (exaltation), 
and, when but slightly virulent, show tendencies to produce in- 
volution forms, as Schron's capsules (attenuation). 

The presence of the pneumococcus in large numbers in a tuber- 
culous sputum would seem to indicate an unfavorable prognosis, 
but the authors have been unable to demonstrate this theory — pos- 
sibly because such a warning usually stimulated vigorous prophy- 
lactic and therapeutic measures. Incubation or centrifugalization 
of a sputum often aids in the finding of the tubercle bacillus. 

Less Frequently Applied Procedures. — These include those ex- 
aminations rarely attempted and also those usually left to the ex- 
pert. In rare instances the physician may desire to search for the 
actinomyces clubs or the echinococcus hooklets. A description of 
these should hardly occupy a place in this work, but the following 
list of less frequently applied procedures is given: 

1. Curschmann's spirals in bronchial asthma. 

2. Heart failure cells in valvular lesions. 

3. Other vegetable parasites, as diplococcus pneumonia, bacillus 
influenzas, and certain molds. 

4. Animal parasites, as ameba coli, etc. 

5. Certain crystals of little or no diagnostic importance. 

6. Albumin tests. 

General Sputum Difficulties. — For those who have difficulty in 
finding Koch 's bacillus, there seems but one remedy — be sure of the 
technic. The correct procedure may be determined by taking a 
sputum known to contain tubercle bacilli and making several 
dozen good stains from it. While making these stains the tech- 
nic will he mastered. It is surprising how expert a physician 
may become after one afternoon's practice and how many hours 
of needless labor he may avoid in the future. 

Some persons do not work with a system, and a negative result 
will be discouraging simply because a repetition of the w r ork means 
another half hour of application to the technic. Apparatus should 
be arranged to save time, and should be kept clean and properly 
classified for use. Several spreads may be made and fixed at one 
time, so that if the first stain fails it will not be necessary to hunt 
up the sample — or possibly the patient — for another test. 



34 LABORATORY METHODS. 

Difficulties in Spreading. — There is a tendency to spread too 
thickly. A droplet of the sputum — not a drop — should be used. 
An attempt should be made to cover every bit of the cover glass 
surface, and a film so thin as to be almost invisible is most likely 
to give the best results. 

Difficulties in Drying. — These difficulties are more serious than 
may appear at first thought, and it is in this part of the technic that 
many failures occur. It is obvious that a thin spread dries much 
quicker than a thick one. Drying should, however, be thorough, 
and it is not sufficient that no water be visible. There may be some 
moisture present — enough to interfere with proper fixation. It is 
a safe rule to wait at least twenty minutes after all moisture has 
apparently disappeared before fixing the specimen. 

Difficulties in Fixation. — These difficulties have been emphasized 
under description of the method of fixation (page 31). Inciner- 
ation, or overfixation, is recognized by the loss of characteristic 
morphology of the spread elements. Underfixation is not impos- 
sible, and is shown by a tendency of the smear not to stain, or, 
when stained, to readily lose the dye. When, after staining, any 
motions of the spread elements are observed, the physician may 
feel certain that the fixation has not been complete. Although the 
proper fixing temperature is learned by experience, a safe rule is 
that the cover glass be hot, but that the preparation be not burned. 

Difficulties in Staining and Washing. — If the stain contains 
sediment, filter it, and, if it seems to be too thin, obtain some fresh 
stain. Although it must steam, ebullition renders the specimen 
unfit for examination because pieces of it are dislodged and float 
off in the stain. Do not allow any portion of the preparation to 
become dry, but add a drop of stain as needed. Two or three 
minutes usually suffice for this procedure. Pour off the excess 
stain and wash immediately in water. Let this washing be 
thorough, so that any precipitated stain may be removed, as such 
particles may be easily mistaken for caseated masses, or even 
microorganisms. 

When the nitric acid causes the stain to fade appreciably, dip 
twice in dilute alcohol and immediately wash briskly in clean 
water. It may be added that one of the most beautiful stains ever 
seen by the authors was made by a physician who, finding that 
his supply of alcohol had become exhausted, substituted gin. 

If a light-blue remains, transfer to fuchsin, but, if the prepara- 



THE SPUTUM. 35 

tion is still dark, wash once more in the nitric acid solution. The 
preparation may not be entirely ruined even if no blue is discerni- 
ble, and it should not be decolorized again merely because a few 
blue lumps occur. These overstained areas indicate thick spread- 
ings, and should not be taken into account at the final examination. 

Difficulties in Mounting. — The slide should be clean. A droplet 
— not a drop — of balsam should be used. Apply the cover glass 
lightly, and do not press it into place. In the colder months the 
balsam may not spread, and, if such be the case, warm the slide 
slightly. If an air bubble clings to the balsam droplet, a pin 
prick will cause its disappearance. 

To Reclaim a Preparation. — Several smears should be spread 
and fixed at one time. If the preparation has been mounted in 
balsam, it may be difficult to put it into a suitable condition for 
restaining, and soaking it in carbol-xylol may aid in removing the 
balsam. One unaccustomed to this work may examine the stains 
in water instead of balsam. 

Value and Limitation of the Sputum Analysis. — Tubercle bacilli 
are often absent in incipient phthisis, but the number of such in- 
stances recorded has been improperly increased by the reports of 
persons who used faulty technic or made wrong observation. 

It is often advisable to substitute for the fuchsin counterstain 
an aqueous picric acid solution. This does not, however, properly 
stain the other microorganisms, but, if the technic has been thor- 
oughly mastered, this disadvantage is more than offset by the ease 
with which the tubercle bacillus may be demonstrated — a bright- 
purple rod on a lemon-yellow background. A deeply stained epi- 
thelial cell to which Koch's bacillus shows a tendency to cling can 
not hide it if this method, as proposed by Mix, is followed. 

The finding of elastic fibers is indicative of "coughed-up" lung, 
providing the vomiting of meat foods has not occurred during the 
coughing up of the sputum, a matter that should be determined. 

It may be accepted as a rule that the finding of acid fast bacilli 
in the sputum indicates pulmonary tuberculosis. Other acid fast 
bacilli are more likely to' be found in pharyngeal secretions than in 
true sputum. The smegma bacillus does not show a curling of the 
ends, nor does it lie in clumps. In case of doubt, other acid fast 
bacilli may be excluded by decolorizing with the alcoholic solution 
of hydrochloric acid described under tuberculosis of the urinary 
organs. (See The Urine in Disease, page 123.) 



36 



LABORATORY METHODS. 




CHAPTER III. 



SEARCHING FOR GERMS. 



Apparatus. — 

1. Aqueous solution of 10-percent 

sodium hydroxid. 

2. Cover glass forceps. 

3. Culture media, prepared and ster- 

ilized. (See page 23.) 

4. Dish of 1:1,000 bichlorid of mer- 

cury solution, colored blue. 

5. Flame. 

G. Gasoline oven designed for one 
burner. 

7. Incubator or substitute. (See 

page 21.) 

8. Microscope and accessories. 

9. Mason jar for unused culture 

tubes. 



10 



thermome- 



Ordinary Fahrenheit 
ter. 

Pan for boiling water. 

Petri dishes. 

Platinum point or loop, or both. 

Slides and cover glasses. 
15. Stains and pipettes — methylene 
blue, Loffler's methylene blue, 
Wright's blood stain. 

Glass tumbler with cotton in bot- 
tom for supporting test tubes, 
platinum wire, pipettes, etc. 

Uterine spoon curet. 



11. 

12. 
13. 

14. 



16 



The arrangement of apparatus is shown in Fig. 7. In this chap- 
ter will be described in a plain, but thorough, manner some of the 
bacteriological examinations which may be made by the country 
physician. Investigations requiring much time, expense, and con- 
siderable skill are not included in these examinations, and animal 
inoculation experiments are avoided. The study of the micro- 
organisms of the bacillus tuberculosis, treponema pallidum, dip- 
lococcus intracellularis, and plasmodium malariae will be found in 
The Sputum (page 32), Vascular Dramas (page 67), Exudates in 
Brief (page 97)., and To Find the Treponema in Six Minutes (page 
173). 

Culture Media. — These may be obtained from certain pharma- 
ceutical houses in assorted tubes of a dozen and properly sterilized 
for use. The following selection should be kept on hand and avail- 
able at a moment 's notice : 



References. — Jordon : General Bacteriology; Stitt: Practical Bacteriology; Novy: 
Laboratory Bacteriology; McFarland: Bacteriology; Muir and Ritchie: Bacteriology. 

37 



38 



LABORATORY METHODS. 



S 



Nutrient agar, four tubes. 

Loffler's serum agar and swabs, four tubes. 

Glucose agar, two tubes. 

Nutrient gelatin, two tubes. 

These media should be kept in a glass jar free from dust, and 
the rubber caps which prevent the ingress of the air should not be 
removed until the media are to be used. After a few months these 
media "dry out" and should then be replaced with new material. 

Influences Which Inhibit Germ Growth. — 

1. Absence or presence of oxygen, depending on whether germ is 
aerobic or anaerobic. The only anaerobic germ considered in this 
book is the tetanus bacillus. 

2. High or low temperatures. The body temperature is most 
favorable to the cultivation of pathogenic microorganisms. 

3. Light. Bacteria work best in the dark, and sunlight is their 
worst enemy. 

Inoculation of Tubes. — "With a bit of the 
material on a sterile platinum wire, inocula- 
tion of suitable media may be accomplished 
by any one of several methods : 

1. Horizontal Streak. The contaminated 
wire is drawn across a plate (Petri dish) of 
culture media. Each germ develops a colony 
where it happens to fall. 

2. Stab. The contaminated wire is pushed 
down the center of solid culture media in a 
tube. Such a method is well adapted for 
anaerobic germs. 

3. Inclined Streak (Fig. 8). This is a 
very convenient method for the rural worker, 
as purchased media in tubes are usually 
slanted. The wire bearing the suspected mi- 
croorganisms is drawn along the center of this 
surface from the bottom upward. The follow- 
ing technic is followed when inoculating by 
this method: 

First. Sterilize the wire in the flame and 
hold with the right hand in such a manner 
that contamination is improbable. 

Second. The tube is held in the left hand 

Fig. 8. — A streak culture. 



SEARCHING FOR GERMS. 



39 



between thumb and index finger, with plugged end toward the 
right. It should be held in a position almost horizontal, so that 
it may not be contaminated by falling dust particles when the plug 
is removed. 

Third. Remove and discard the rubber cap. The cotton plug- 
is grasped between the middle and third fingers of the right hand 
and quickly removed with a sharp twist. 




Fig. 9. — Correct method of inoculating a tube. 



Fourth. Hold the neck of the tube in the flame for a moment 
in order to incinerate any microorganisms which may be attempt- 
ing to gain entrance. 

Fifth. Now inoculate the tip of the sterile wire with the 
smallest bit of the suspected material and streak the agar as de- 
scribed above, being careful not to contaminate anything (Fig. 9). 
Especially avoid air currents, and do not allow the platinum point 
to touch the side of the tube. 

Sixth. Withdraw wire. 

Seventh. Again pass neck of tube through flame. 

Eighth. Insert cotton plug. 



40 LABORATORY METHODS. 

Ninth. Sterilize wire in flame. 

The entire technic, when properly learned, should take only 
about one minute, and should be conducted with care. The in- 
oculated tube should be kept at body temperature for twenty-four 
hours. 

Small colonies should mark the path of the wire. Unless the 
least amount of the suspected material has been used, these will, 
because of their great number, coalesce and render isolation of 
the causative germ impossible. A platinum point is, therefore, 
more desirable than a loop. Separation, if necessary, must be done 
at once, as overcrowding as well as the presence of involution 
forms will occur as time passes. 

Isolation of Pure Cultures. — Specimens from the various colo- 
nies may be examined at once, and this is usually an advisable 
procedure in diagnostic work. It may, however, be the desire of 
the worker to obtain a pure culture of the microorganism — i. e., 
where the tube contains only this germ and coalescence of colonies 
does not cause contamination — which may be done as follows: 

First. Place several tubes of nutrient agar upright in a can of 
boiling water. When liquefaction of the media, has occurred, per- 
mit the water to cool down to about 122° F. This is the inoculat- 
ing temperature, solidification occurring as cooling progresses be- 
low this point. 

Second. Meanwhile, with a hand lens, study the colonies on 
the original slanted surface. Some may be round and others 
irregular, some transparent and others opaque, and some flat and 
others raised. A certain germ invariably gives rise, in its growth, 
to a colony typical of its species, and inoculations from a single 
colony should result in a pure culture of that germ. In case all 
colonies appear alike, the primary inoculation was possibly a pure 
culture. 

Third. When the melted agar reaches 122° F., inoculations 
should be made at once according to the method described above. 
In this case a platinum point, rather than a loop, should be dipped 
into the colony, as otherwise too many microorganisms will be ob- 
tained. Each inoculation should be mixed well into the agar, and 
the wire resterilized in the flame before inoculating the next tube. 

Fourth. The liquid agar, thus inoculated, is immediately (after 
heating neck of tube) poured into a Petri dish that has been 
previously heat sterilized. The tube is then thrown into a strong 



SEARCHING FOR GERMS. 41 

solution of mercuric chlorid and later cleansed by boiling. The 
Petri dish is immediately, with its contents (plate preparation), 
set aside at blood heat. Other dishes are inoculated in the same 
way. 

The above technic requires haste, as the agar cools rapidly and 
solidifies. 

Authors' Short Method. — The authors are convinced that, for 
practical purposes, the above plating is not always necessary. The 
process is tedious, and, though usually described as the standard 
method, offers so many opportunities for contamination that the 
practitioner is justified in seeking further for a substitute. 

In this book the statement has been emphasized that, although 
isolation of pure cultures is not a necessary procedure in many 
diagnostic problems, the worker soon takes pride in this separation. 
If, however, such isolation is avoided, an early examination of 
specimens from the colonies is often imperative for the reason that 
coalescence of colonies and involution forms of the germs occur. 
A simple method of isolating pure cultures is to select samples 
from the various colonies and inoculate other agar slants. When 
this Avork is done early and carefully, it will not be subject to criti- 
cism. 

Incubation in Vacuum Bottles. — Reference has been made to 
substitutes for the expensive and complex sterilizer and incubator. 
If a so-called vacuum bottle or a tireless cooker be employed, it 
will be necessary to see that the cultures are obtaining a proper 
amount of oxygen. Such an incubator should be kept in a warm 
room, and the cork or lid inserted only after both the culture and 
the inclosed air are warm. The required warmth may be secured 
with a slightly heated stove. It may be advisable to have this 
temperature a little higher than 98.6° F., as some cooling will 
always occur. In order to insure -plenty of oxygen, it is best to 
"reincubate" at least once during the twelve hours or twice dur- 
ing the twenty-four hours. A little sweet oil heated to body 
temperature may be placed in the bottom of the vacuum bottle, 
but plenty of air must be left above the surface for the use of the 
developing colonies. 

Sterilization. — 1. Nutrient Media. These should be purchased 
ready for use. 

2. Glassware and Metalware. These should be boiled for at 
least ten minutes, making sure that all portions are touched by the 



42 LABORATORY METHODS. 

water, after which they are cleaned and dried. Contamination 
with various atmospheric bacteria occurs during the cleaning, so 
that Petri dishes and other glassware treated in this manner are 
not available for further culture work without hot air sterilization 
just before using. The sterilization may be conducted in a gaso- 
line oven, provided that sufficient time and care be taken. The 
degree of heat required — about 300° F. — necessitates removal of 
all cotton plugs and organic material. Glassware, but not metal- 
ware, may be immersed in a strong solution of mercuric chlorid 
before boiling, which does not usually kill the germs, but weakens 
them to such an extent that they easily succumb to the boiling 
water. 

3. Cover Glasses. These, when cleaned and dried, may be 
passed through the flame several times preliminary to making a 
spread, and may be heated past the fixing point, so that all organic 
material is destroyed. After cooling, they are practically sterile 
and the spread may be made at once. 

4. Towels. These may be dipped into the mercury solution, 
but a thorough boiling should never be omitted. During the boil- 
ing every portion should be immersed in the water. 

5. Hands. During culture work the hands should not come in 
direct contact with any sterile apparatus, nor with any suspected 
virulent material. The fingers should not touch the platinum 
loop, lower end of the cotton plug, inside of the tubes, or Petri 
dishes known to be sterile, and should not, of course, come in con- 
tact with colonies, pus, etc. At frequent intervals the hands 
should be scoured with a brush and hot suds. Unless mercuric 
chlorid causes dermatitis in the worker, he may keep a 1 : 1,000 
solution conveniently near, into which to dip his hands occasionally, 
and thus inhibit the effect of any germ which might alight at the 
mouth of a sweat gland. For convenience of distinction, a little 
blue or green coloring matter may be added to this solution. The 
physician who looks on these solutions not as antiseptics, but as 
disinfectants, has something to learn. 

6. Metal Instruments. Except for the mercury solution, 
which ruins them, these may be treated in the same manner as 
glass. Forceps, wires, etc., may be heated to incandescence in the 
nonluminous flame. 

7. Laboratory Tables. Should be wiped off at frequent inter- 
vals with a cloth saturated in 1 : 500 solution of mercury. 



SEARCHING FOR GERMS. 43 

Simple Stains. — Of these the authors prefer methylene blue in 
most diagnostic procedures, as this dye usually stains well, but 
seldom overstains. Other good stains are fuchsin and gentian 
violet. The methylene blue stain is prepared by diluting some of 
the concentrated alcoholic solution with water, the object being to 
obtain a stain with little alcohol. It is best, however, to dilute 
slowly until the liquid is transparent, as otherwise precipitation 
of the stain will occur on the cover glass, and, if this happens, re- 
ject the stain and make up some more, using less water. An un- 
saturated stock solution gives, when diluted, a weak stain, and 
hence the necessity of obtaining these dyes from a reliable firm. 
The following technic for staining bacteriological smears will give 
excellent results: 

First. Spread thoroughly some of the material on a sterile 
cover glass with a sterile platinum loop. This wire is always' ster- 
ilized by heating to incandescence in the nonluminous flame. 
Remember that a spread is rarely or never too thin, but is often 
too thick, or at least may be so in spots. 

Second. Dry in air. 

Third. Fix in flame according to directions given under sputum 
(page 30). 

Fourth. Permit preparation to cool. 

Fifth. Grasp the glass with forceps and lock, and, holding 
specimen side up, add enough stain to cover the smear. 

Sixth. After about one minute — exact time depending on 
strength of stain — wash off the dye under the tap or dip repeat- 
edly in a bowl of water. 

Seventh. Dry lower surface of glass with a piece of filter paper 
or an ordinary blotter, invert, and float, specimen side downward, 
onto a clean slide. 

Eighth. This serves as a diagnostic examination. In case it is 
desired to keep the preparation, it may be dried between two blot- 
ters and mounted in a droplet of balsam (page 34). 

Staining Errors. — Overstaining with methylene blue is rare. 
Understaining, however, is common, and may depend on one of four 
causes — overfixing, underfixing, a weak staining fluid, or insuffi- 
cient time of exposure. Overfixation is usually apparent by the 
injury to the bacteria — charring, etc, Underfixation permits the 
floating off of the specimen during washing. Underfixation may 
be microscopically demonstrated by the motility of the germs, even 



44 LABORATORY METHODS. 

though partially stained. In case staining troubles should con- 
tinue, it is possible that the solution is too dilute, and precipita- 
tion of the dye may indicate this condition. In case the staining 
time was insufficient, the preparation should take the dye on the 
second trial. 

Other Staining Methods. — There are many excellent stains in- 
dicated in other books, but which can hardly be recommended in 
this work as absolutely necessary for the diagnostician, who must 
reserve many of his energies for other kinds of labor. In searches 
for the gonococcus and diphtheria bacillus, the physician may 
advantageously substitute Loffler's methylene blue. Gram's dou- 
ble stain, spore stains, flagella and capsule stains — and indeed 
many others not mentioned in this chapter — should be understood 
by the practitioner in order that the value and limitations of all 
bacteriological work may be borne in mind. 




Fig. 10. — Surface colonies of diphtheria "bacillus. These become quite characteristic 
within eighteen hours, while the other microorganisms may not yet be visible. The 
colony is flat, with a wavy edge and a dark center. The contents are grayish white, 
coarsely granular, and not unlike ground glass. Drawn from an eighteen-hour cul- 
ture. 

Searching for the Bacillus Diphtheriae. — At least one physician 
in the radius of every one hundred miles should be prepared for 
this examination, and no county seat should be without a person 
equipped for this work. The procedure is not difficult, and, if 
the examination is conducted by the physician in charge of the 
case, it will be more valuable than if made by a person — often not 
a physician — several hundred miles distant. 

A swab from the throat may be tested immediately, as smears 
can be made, stained, and examined at once. To be certain of the 
diagnosis, an inoculation may be made which, within eighteen 
hours, should show hundreds of the Klebs-Loffler bacilli. Isolation 
in pure cultures of this microorganism is not necessary for the 



SEARCHING FOR GERMS. 45 

diagnosis of diphtheria. The serum agar slant tubes and swabs 
may be purchased with other media. No disinfecting solutions 
are to be used on the throat preliminary to the swabbing, for which 
the procedure is as follows : 

1. Rub swab over affected portion of the throat, removing, if 
possible, some of the membrane. 

2. Streak over surface of serum agar. Several such tubes should 
be so prepared. 

3. Keep at blood temperature from eighteen to twenty-four 
hours. 

Diphtheria Colony. — This is moist in appearance, large, round, 
and of grayish color (Fig. 10). The center is thick, while the 
edges are thin, and examination with a hand lens shows a wavy 
border. Cover glass smears may be made from these colonies, 
fixed, and stained Avith Loffler 's methylene blue. 






Fig. 11. — Some of the more common forms of the diphtheria bacillus, reproduced from 

actual preparations. 

Bacillus Diphtheriae. — The Klebs-Lomer bacillus shows some or 
all of the following characteristics as compared with other bacilli: 

1. Irregularity of form and size, illustrated in Fig. 11. 

2. Club shapes, or bacilli bearing swollen ends. 

3. Cross striations or bands. 

4. Monopolar or bipolar staining. 

5. Wedge shapes. 

6. Curved forms. 

Certain forms of bacilli — pseudo types — may closely resemble 
the true bacillus of diphtheria. In case of doubt their character 
can be determined by allowing them to come in contact with a drop 
of water, when the pseudo type will form a cloudy suspension, 
which will not occur with the true Klebs-Lomer bacillus. 

Searching for tho Bacillus Tetani. — The cultivation of the 
tetanus bacillus requires the absence of oxygen. Many methods 



46 



LABORATORY METHODS. 



have been devised, but the method given here is suggested, and 
smears from the wound may be examined. It is a well-known fact 
that anaerobic germs may multiply in the presence of those requir- 
ing oxygen, a phenomenon that is termed "microbic association," 
and therefore, if any tetanus bacilli in the wound are capable of 
multiplying — unless planted very deeply in the tissues — the pres- 
ence of certain oxygen-requiring germs is presumed. 

1. With a sterile uterine spoon curet scrape well the wound, 
being certain that no antiseptic has been used previous to this 
procedure. A thorough cauterization for therapeutic purposes 
should follow this curettage. 

2. Add some of this material to a tube of serum agar — not a 
slant, but one which has been previously liquefied, if necessary, and 
resolidified in a vertical position (page 40). 

3. Incubate from four to seven days. 

4. Examine for characteristic drum-sticks, which are not easily 
found, for, even if the culture be pure, all tetanus bacilli do not 
show the end spore, and typical drum-sticks often escape detection 
because simple stains do not bring out well the end spore. Hot 
carbol-fuchsin will stain both the spore and the body. (For 

method see page 31.) The tetanus bacillus, 
with or without a spore, is usually a very 
long and slender rod. 

5. Heat the culture in water up to 175° F. 
and maintain at this temperature for forty- 
five minutes. Only spores remain viable. 

6. After cooling, make a stab culture in 
glucose agar and incubate. Such tetanus 
spores as may be present will develop deep in 
the agar and show a cloudy growth, with 
characteristic perpendicular branchings. Its 
indistinctness, together with the fir tree ap- 
pearance, is peculiar to the microorganism. 
The gelatin stab culture is much more distinct 
(Fig. 12). 

Sources of Error. — The isolation of the tet- 
anus bacillus is not an encouraging proce- 
dure, even for the expert. Strange to note, 
however, the authors have seen it identified by persons who would 
be least expected to find it. The attempt is worth the effort, 




Fig. 12. — Typical fir tree 
tetanus stab in gela- 
tin. Six days' cul- 
ture. Occurs only in 
the lowest portions 
of the inoculation. 



SEARCHING FOR GERMS. 47 

although the spores of other anaerobes may interfere. Stitt at- 
taches importance to the odors which may arise from the serum 
culture, and states: "From day to day smell the culture, and, if 
an odor similar to the penetrating sour, foul smell of the stools 
of a man who has been on a debauch is detected, it is suspicious. 
The nondevelopment of a foul odor is against tetanus." 

Searching for Koch- Weeks Bacillus. — This may be necessary 
when attempting to differentiate a severe catarrhal conjunctivitis 
from a mild gonorrheal infection. Smears from the secretion 
should be made and stained for the gonococcus (page 48). If the 
Koch-AYeeks microorganisms, instead of the diplococci, are present, 
they will appear as very short bacilli. If the gonococci are present, 
they usually, though not invariably, lie within the protoplasm of 
the pus cells. 

Searching for Streptococcus Pyogenes. — This important micro- 
coccus has been found in the following conditions : 

1. Diffuse inflammations, as phlegmons, erysipelas, etc. 

2. Inflammations of respiratory tract, as pharyngitis, bronchitis, 
lobular pneumonia, etc. 

3. Acute articular rheumatism. 

4. Ulcerative endocarditis. 

5. Puerperal fever. 

6. Scarlet fever. 

7. Certain inflammations of serous linings, as peritonitis, etc. 

Some authorities claim that the streptococcus pyogenes is a dis- 
tinct microorganism, being closely related to other chain cocci, but 
not identical with those causing erysipelas, rheumatic fever, and 
scarlet fever. So far as diagnostic purposes are concerned, this 
distinction is unimportant. 

The streptococci occur normally in the mouth, nasal cavity, large 
intestine, vagina, and on the integument. The attenuation and 
exaltation of the streptococcus seems an easy matter, and the 
resisting powers of the organism need suffer only slight decrease 
to render it an easy prey. In pus, search first for the staphylococ- 
cus and then for the streptococcus, but follow vice versa in the 
diffuse and more extended inflammations. The isolation and identi- 
fication of this coccus by cultural examination is hardly to be 
recommended to the country physician, as there are so many sources 
of error and so many limitations. Smears stained with methylene 
blue offer a much better, though still imperfect, method. Usually, 



48 LABORATORY METHODS. 

though not invariably, the long chains of streptococci, compared 
with the short chains of less than eight cocci, are the really dan- 
gerous forms, but it has been proven that virulent short chains do 
exist. 

Searching for Staphylococci. — This is the most common of all 
pus germs, and for practical purposes need not be divided into 
subclasses. It is, however, interesting to note that furuncles and 
other abscesses of the integument are due usually to the varieties 
which form yellow colonies, while those of pharyngeal infections 
are white. Suspected pus may be inoculated into nutrient agar by 
following the directions applying to diphtheria. The opaque 
creamy-white or light-yellow colonies are most likely to show the 
staphylococcus. Both forms liquefy gelatin, and often the germ 
may be found in the pus smears. 

While methylene blue serves well as a stain, the use of Wright's 
blood stain, when much work is being done with pus, is specially 
recommended as an occasional substitute for methylene blue, for, if 
one stain is misplaced or spilled, the worker will not be forced to 
discontinue the examination. Wright's stain, though requiring 
distilled water, does not make heat fixation necessary. 

Isolation of the Typhoid Bacillus. — With our present knowledge 
the practitioner should be content to leave this work to specialists. 
At best it is tedious, and, when attempted from suspected water, 
is usually a very discouraging procedure. 

Pneumonia Versus Tuberculosis. — "Galloping consumption" 
may resemble lobar pneumonia, and the differentiation of these 
conditions- is considered under sputum analysis. The pneumo- 
coccus is very small and is seen only with the highest powers. 
The capsules may best be demonstrated in hanging drop prepa- 
rations. Cultures are unsatisfactory. 

Searching fcr the Gonococcus. — No cultures should be at- 
tempted. Loffler's methylene blue stains well, as does also Wright's 
blood stain. The typical form of this coccus, its occurrence in 
pairs, and its tendency to lie within the pus cells are its character- 
istics, and it is not necessary to use Gram's stain in order to avoid 
confusion with other micrococci. 

Oil immersion objective should be used. Three drams of gin, 
taken twelve hours before the examination, usually drives this 
germ out of its hiding places in the gland ducts. 

Searching for the Colon Bacillus. — A germ which, once con- 



SEARCHING FOR GERMS. J | 

sidered harmless, now lias the distinction of chief etiological factor 
in the following conditions : 

1. Some cases of cystitis. 

2. Some cases of peritonitis, especially those resulting from per- 
foration of the bowel or appendix. 

3. Some cases of appendicitis. 

4. Seme cases of mucous colitis. 

The bacillus coli communis is a normal and. doubtless, a neces- 
sary inhabitant of the large bowel. Neither the morphological nor 
the cultural properties of this microorganism are sufficiently char- 
acteristic to justify its search. The presence of rods in urine does 
not, as is commonly supposed, warrant the diagnosis of colon 
cystitis. For its relation to sewage contamination see Some Sim- 
ple Water Analyses, page 145. 

Searching for Molds. — There are three very common pathogenic 
molds : 

1. Microsporon furfur of fawn chest, or tinea versicolor. Scrap- 
ings from the skin should show, besides epidermal cells, the hyphae 
of this mold, and intermingled are loose spores which show a 
tendency to clump formation. 

2. Endomyces albicans of thrush; segmented hyphaa, with spore 
formation within ; and branchings may be seen at segments. 

3. Trichophyton of barber's itch, ringworm, etc.; a number of 
varieties showing hyphas. 

Therapeutic indications make an absolute classification of these 
varieties for the practitioner unnecessary, and it is sufficient to 
find hyphas or spores, or both. The following technic has given 
excellent results : 

1. Make scrapings from the edge of the lesion, and never from 
the central portion. 

2. Drop these on a clean slide, and add 2 or 3 drops of 10-percent 
sodium or potassium hydrate solution. 

3. Apply lightly a clean round cover glass and let stand fifteen 
minutes. 

4. Firmly press down the cover glass, flattening the specimen. 

5. Examine with a somewhat contracted diaphragm and a high- 
power objective. 

Searching for Actinomyces. — Besides the ordinary "jaw infec- 
tions." cases of consumption, ischiorectal abscess, and furunculosis 
have been observed in which the chief etiological agent seemed to 



50 LABORATORY METHODS. 

be the ray fungus. Cultivation for diagnostic purposes is hardly- 
worth while. The characteristic yellowish granules may often be 
observed in the pus without the aid of a lens. Smears may demon- 
strate the characteristic clubs; if they do not, curettage of the 
walls of the abscess may loosen the fungus. 

Other Searches Which May be Attempted by Smears Alone. — 
Culture work is unnecessary in many bacteriological examinations. 
Many of these have been included in other chapters — viz., trepo- 
nema, tubercle bacillus, Plasmodium, and meningococcus. Others 
may be attempted by the smear method, as : 

1. Spirocheta of Vincent's angina; in certain throat ulcers. 

2. Ameba coli and cholera vibrio. (See Every-Day Stool Tests, 
page 150. 

3. Bacillus influenzas from greenish expectoration. 

4. Bacillus pyocyaneus from green pus. 

Researches. — All medical truths have not originated in medical 
schools. or hospitals, and there is no reason why a general practi- 
tioner should not have discovered the treponema pallidum just as 
a country doctor first identified the germ of tuberculosis.^ As an 
illustration of what is yet to be discovered, the following list of 
diseases — of an infectious character, but of doubtful or unknown 
etiology — are submitted to the country physician for his consider- 
ation, of which diseases the leaders of scientific research have thus 
far failed to find the cause, although much expense has been in- 
curred by investigations: 

1. Infantile paralysis. 

2. Pellagra. 

3. Rabies. 

4. Acute articular rheumatism. 

5. Measles. 

6. Mumps. 

7. Scarlet fever. 

8. Smallpox. 

9. Whooping-cough. 
10. So-called colds. 

Difficulties and How to Avoid Them. — Many sources of error 
are self-evident, and need no consideration, while some errors are 
made in following the various methods. If, however, the technic 
of culture work is closely followed, there should be very little 
trouble. 



SEAECHING FOR GERMS. 51 

Value and Limitation of These Searches. — Both value and limi- 
tation are variable, not only as to the various microorganisms, but 
also as to the physician — that is, a procedure which may be of value 
in the hands of one person will be a waste of time in the hands of 
another. A physician who has not finished a course in laboratory 
bacteriology should be slow to handle the more virulent germs, and, 
on the other hand, the late graduate may attempt searches not 
recommended in this book. A safe rule will be, before reaching 
any conclusion as to the identity of any microorganism or its re- 
lation to the pathological process at hand, to thoroughly study 
the subject from the best works on bacteriology, as in this book 
only those examinations have been selected and classified which may 
be profitably employed by the general practitioner. 

Searches Left to Experts. — In the light of our present knowl- 
edge of this relatively new subject, it appears that the practitioner 
may wisely avoid working with some germs, as follows: 

1. Germs identified with difficulty: colon bacillus, typhoid bacil- 
lus, paratyphoid bacillus, germs of food poisonings, dysentery ba- 
cillus, tetanus bacillus. 

2. Germs exceedingly virulent : plague bacillus. 

3. Germs rarely found : anthrax bacillus. 

Laboratory Prophylaxis. — For additional information on this 
subject see Laboratory Prophylaxis, page 179. 



52 



LABORATORY METHODS. 




CHAPTEE IV. 



VASCULAR DRAMAS. 



Apparatus. — 






1. Acetic acid, glacial. 


13. 


Slides and cover glasses. 


2. Acetic acid, 1-percent. 


14. 


Slide forceps. 


3. Alcohol. 


15. 


Sodium chlorid, aqueous solution, 


4. Broom straw. 




.85-percent. 


5. Distilled water. 


16. 


Sticker or stub pen. 


6. Ether. 


17. 


Stiff paper. 


7. Glycerin. 


18. 


Tallqvist hemoglobin chart. 


8. Hayem's solution. 


19. 


Towel. 


9. Horse hairs. 


20. 


Wright's blood stain; in 1-ounce 


10. Marx's fluid. 




bottles in liquid form from 


11. Microscope and accessories. 




some reliable chemical or op- 


12. Red and white pipettes and count- 




tical firm. 


ing chamber. 







The arrangement of apparatus is shown in Fig. 13. With a 
little practice any physician may become expert in the vari- 
ous blood analyses. While some examinations are of importance, 
they must not be overestimated, as a diagnostic chain usually re- 
quires many strong links before it becomes useful to the thera- 
peutist, and only the few most valuable procedures are recom- 
mended. One method, the best and probably the most simple, is 
described briefly, but no necessary details are omitted, and sources 
of error are pointed out unless self-evident. The actual selection 
of the necessary examinations for a given case lies with the physi- 
cian, it being hardly worth while to make complete blood examina- 
tions in every case. In this selection the diagnostician must not 
go astray, and it is unwise, for example, to conclude that, inasmuch 
as a low hemoglobin is observed, the red count would be diminished 
and the counting of the erythrocytes neglected. 

Development of the Blood Cells. — It is probable that all blood 
cells, red or white, normal or pathological, originate from one 



References. — Ewing: Pathology of the Blood: Schleip : Atlas of Hematology: Cabot: 
The Blood; Da Costa: Clinical Hematology; Watkins: Diagnosis by the Blood; all 
works on clinical diagnosis, including Sahlij Simon, Boston, Wood, Emerson, Webster, 
etc. 

53 



54 



LABORATORY METHODS. 



parent cell, and, with the exception of many of the lymphocytes, 
all blood corpuscles are formed in the red bone marrow. The 
spleen, once termed the "cradle and the grave" of the blood, by 
no means fills either office, but may serve often as a brooder for 
the younger corpuscles and as a subheart — a "portal pump," 




Fig. 14. — Development of the blood cells. Schematic representation of the corpuscles 
normally found in the red bone marrow, showing how they give rise to the various 
circulatory elements. Cells normally present in bone marrow: 1, parent cell; 2, in- 
termediate forms, with neutrophilic, basophilic, and eosinophilic alterations; 3, neu- 
trophilic myelocyte; 4, basophilic myelocyte; 5, eosinophilic myelocyte; 6, erythro- 
blast. Cells normally present in circulation: 7, neutrophilic leukocyte; 8, basophilic 
leukocyte; 9, eosinophilic leukocyte; 10, lymphocyte; 11, transitional; 12, erythro- 
cyte; 13, blood plate. 



which, by the slow contractions of its muscular walls, forces the 
blood through the liver to the central pumping station. Hepatic 
rather than splenic tissue forms the tomb for the erythrocytes. 

A modified form of certain diagrams offered by Schleip, with 
nomenclature according to the suggestion of Grawitz, is given in 
Fig. 14. 



VASCULAR DRAMAS. 55 

Blood Dramas. — For convenience of description, we may con- 
sider each blood picture a "drama," the circulatory system the 
"stage," and the red bone marrow the "back of the scenes." In 
the normal blood picture the various circulating corpuscles are of 
constant variety and in practically unvarying number, and in 
differential counts there are no marked percentage alterations. In 
disease, however, the scene is changed and there is turmoil at once. 
In these conflicts certain types may be deformed or destroyed, 
others may be increased in number, and finally, in the so-called 
crises, those cells not ordinarily observed in the circulating fluid 
may be called from the home or the brooder to aid their struggling 
offspring. It must, therefore, be concluded that the so-called blood 
diseases do not give rise to corpuscles hitherto not found in the 
circulating blood or in the blood-forming organs. Every morbid 
blood scene has, for its characters, dead, dying, or wounded normal 
cells along with certain other varieties in increased numbers — 
active or injured maternal reinforcements. By keeping these 
points in mind, the significance of each blood smear can be realized 
without difficulty, and the pathology of blood diseases will be easily 
comprehended by the clinician. 

Behind the Scenes. — Here is the busy spot. For example, in 
pernicious anemia, as the red cells are destroyed, there is call for 
reinforcements. The organism demands even more oxygen than 
usual, and an extra strain is put on the bone marrow. More and 
more red cells are rushed into the circulation, but to no avail. 
Finally the bone marrow becomes fatigued — if we may use such 
term — as those storehouses in the spleen, hemolymph glands, and 
perhaps other organs and tissues have been exhausted, and the 
demand exceeds the supply. The result is an inferior product, 
or, to adhere more closely to our comparison, "understudies." 
Cells, hardly fitted for the assumption of the duties thrust upon 
them, are rushed to the fray. Among these are found the over- 
sized and undersized cells, forms showing mitoses and red cells 
which have not yet cast aside their nuclei. They are quickly 
destroyed, and their dying forms — polychromophilia, poikiloblasts, 
etc. — must not be mistaken for cells entirely foreign to the organ- 
ism. Then the bone marrow rouses and concentrates its energies 
in one mighty effort ; there results the blood crisis, the sending out 
of certain forms not only unfitted by improper development for 
the struggle, but often already diseased — the megaloblasts. In 



56 * LABORATORY METHODS. 

this manner a little study may make plain the picture or scenes of 
any blood disease. 

Cast. — The complete description of blood corpuscles will be 
found in the larger books, but a few brief hints may not be amiss. 
In all cases where the colors of stained preparations are considered 
it is to be understood that reference is made to Wright's stain. 
Although blood cells appear flat when spread, fixed, and stained, 
such is not their appearance in the natural state. All white cells 
are globular, the polymorphonuclear forms showing pseudopods. 
The red cell is not a biconcave disk, as was the former conception, 
but is thimble-shaped — i. e., one surface is very convex and the 
other deeply concave, such morphology being a direct result of the 
extrusion of the nucleus of the erythroblast. These nuclei, in turn, 
are supposed to give rise to the blood platelets. 

For convenience, the blood corpuscles have been arranged in sys- 
tematic order, and only those characteristics observed in the stained 
preparation are considered : 

Red Cells.— 

1. Erythrocyte; the normal red cell; orange or pink. 

2. Undersized cell; sometimes normal. 

3. Microcyte; very small erythrocyte; invariably pathological. 

4. Oversized cell ; sometimes normal ; in large numbers are char- 
acteristic of pernicious anemia. 

5. Macrocyte or megalocyte ; very large red cell ; this and re- 
maining forms of red cells are always pathological. 

6. Poikilocyte; irregular form, with beak or snout. 

7. Shadow or ghost ; loss of hemoglobin, with distended cell wall. 

8. Endoglobular degenerations; appear as unstained droplets 
within the protoplasm. 

9. Enlarged delta. 

10. Small delta. 

11. Normoblast; with single deep-blue nucleus. 

12. Microblast; small normoblast. 

13. Macroblast or megaloblast ; macrocyte, with pale nucleus and 
signs of degeneration. 

14. Poikiloblasts ; nucleated poikilocytes. 

15. Degenerating normoblasts; showing lobulated nuclei. 

16. Forms showing karyorrhexis ; granular disintegration of 
nuclei. 

17. Forms showing karvokinesis : division figures in nuclei. 



VASCULAR DRAMAS. 57 

18. Polychromatophilia or paradoxical stain; where nuclear dust 
of karyorrhexis mixes with hemoglobin, causing a bluish tint. 

White Cells. — Percentage refers to the differential count in 
adult's blood. 

1. Lymphocyte (19 percent) ; robin 's-egg-blue cytoplasm; round 
purple nucleus. 

2. Transitional (3 percent) ; light-blue cytoplasm; horseshoe 
nucleus. 

3. Polymorphonuclear eosinophile (2 percent) ; cytoplasm con- 
tains large and deep-red granules ; lobulated nucleus. 

4. Polymorphonuclear basophile (1 percent) ; cytoplasm contains 
medium-sized, light-blue granules ; lobulated nucleus. 

5. Polymorphonuclear neutrophile (75 percent) ; cytoplasm con- 
tains small reddish-lilac granules; lobulated nucleus. 

6. Myelocytes containing the various colored granules ; these are 
very large cells, with single nuclei; pathological. 

7. Degenerating leukocytes ; cytoplasm contains vacuoles ; patho- 
logical. 

8. Disintegrating leukocytes; disappearance of cytoplasm, with 
a scattering of the granules ; pathological. 

9. Necrotic changes; nuclei stain poorly or not at all, or else 
show fragmentation. 

Blood Dramas. — The table given on page 58 serves to differen- 
tiate the more common blood conditions. For detailed information 
reference should be made to the larger books. 

Bedside Apparatus. In case the patient is bedfast, suitable 
specimens of the blood may be obtained and taken to the office for 
examination. Dilutions and spreads may be completed with little 
apparatus, which may be easily carried in the physician's hand- 
bag, as: 

T. Towel. I 7. Vial containing (1-percent) acetic 



2. Blood sticker or stub pen. 

3. Hemoglobin chart. 

4. Red and white pipettes. 

5. Catheter tubing. 

6. Vial containing Hayem's solution 

for reds. 



acid for whites. 
Vial containing alcohol. 
Six clean slides and some stiff 

wrapping paper. 



The towel should "be clean, and a clean towel or handkerchief 
can usually be obtained at the home of the patient. A nickel- 
plated German blood sticker offers no advantage over a stub pen 



58 



LABORATORY METHODS. 



with one point broken off. Arneill recommends small rubber 
catheters instead of the tubing supplied with blood pipettes, and, 
if these are sufficiently long, the free end may be brought around 
ana slipped over the point, in which condition the diluted and 
prepared blood may be transferred to the office and examined at 
leisure. A blood spread which has dried may be wrapped in heavy 
paper and taken to the office. 

THE MORE COMMON BLOOD CONDITIONS. 1 



Disease 

or 

condition 


Red 

cells 


Hemo- 
globin 


White 
cells 

N 


Chlorosis 


N 


— 


Polycythemia 


+ 


Xor + 


N 


Secondary ane 








mia 






X or — 


Pernicious or 
large-celled 
anemia 


— 


— 


— 


Leukopenia 


N or — 


X or — 


— 


Leukocytosis 


N or — 


N or — 


+ 


Leukemia 


— 


— 


+ 


Eosinophilia 


— 


— 


+ 


Hodgkin's dis- 
ease 

Lymphosarco- 
ma 


— 


: 


+ 


Iodophilia 


N 


X 


+ 



Cytology 



(a) Post-hemorrhagic — undersized red cells, 
few poikilocytes, some endoglobular de- 
generation, normoblasts, (b) Cachectic or 
toxic — microcytes, undersized cells, more 
poikilocytes, more degeneration; normal 
and degenerating normoblasts. 

Loss of hemoglobin greater than above, mod- 
erate degeneration, oversized cells, macro- 
cytes, megaloblasts, polychromatophilia, mi 
toses, degenerating normoblasts, less poi- 
kilocytosis than above. 

A condition in typhoid, acute miliary tuber- 
culosis and pernicious anemia; white 
count decreased due mainly to diminution 
in the polymorphonuclears; lymphocytes 
are increased. 

Xoted in purulent infections; polymorphonu- 
clears actually increased; lymphocytes rela- 
tively, but not actually, diminished. 

(a) Lymphatic — increase of the lymphocytes, 
(b) Myelogenous — increase of the poly- 
morphonuclears, appearance of myelocytes 
in the blood. 

Increase of eosinophilic polymorphonuclears; 
noted in trichinosis, asthma, uncinariasis, 
menstruation, positive tuberculin, certain 
skin diseases, etc. 

Picture of a cachectic secondary anemia. 

Large cells or cells of various sizes, re- 
sembling very much the lymphocyte, but 
staining poorly or showing degenerations. 

Certain iodin reaction seen in the polymor- 
phonuclears when purulent processes are 
present in the body. 



Obtaining the Blood. — Although the lobe of the ear furnishes 
blood in considerable quantity, it may be often necessary in bed- 
fast patients to use the finger tip. A cold, bloodless integument 
does not represent that actual blood picture which may be present 
at other parts. If, however, a massage is undertaken, it is best to 



X, normal; +, increased; — , diminished. 



VASCULAR DRAMAS. 59 

wait until the deep-red color is replaced by the normal tint. The 
puncture should be made quickly, and should be fairly deep, so 
that repetition will not be necessary. 

Macroscopical Examination. — Except in aggravated conditions, 
such an examination is of little use in diagnosis. A mushy blood 
may be due to a severe leukemia, a "whey" solution may suggest 
a great decrease in the number of cells, a pale blood signifies hemo- 
globinemia, and chocolate or crimson tints may be due to certain 
poisons. Without a suitable chart, colors are often misleading, and 
the laity watch with interest the changes in the color of the blood 
when in reality tests would show that there is no alteration in the 
hemoglobin. 

Microscopical Examination of Fresh Blood. — Experts are often 
able to pass a final conclusion from a mere inspection of a hanging 
drop. Although the practitioner may make certain interesting- 
observations, no final diagnostic inferences should be drawn from 
this procedure, and, if time is of importance, he may well omit this 
examination. 

Hemoglobin Estimation. — For practical purposes, the Tallqvist 
color chart is sufficient. The original booklet form has a decided 
advantage over its modifications, as it is convenient to carry and is 
not easily soiled. Instructions accompany each booklet. The 
comparison of the color of the droplet with that of the chart should 
be made at once, as certain changes take place, on drying, in the 
hemoglobin. In case the physician has no hemoglobin chart with 
him, he will not be entirely at a loss, as comparison of a drop of the 
patient 's blood with his own on a piece of white blotting paper may 
bring out good color distinctions. 

Suspension of Red Cells. — Blood, after coagulating in pipettes, 
gives considerable trouble, and the beginner may find it advan- 
tageous to practice with a little carmine water. Dilutions must be 
made quickly. Wipe away any blood which may be present at the 
puncture, and, when the next drop appears, immerse well the 
point of the red pipette below its surface and draw, by mouth 
suction, a solid column of the blood to the mark 0.5. Quickly 
wipe off the point and instantly immerse it into the vial of Hayem's 
solution. Now quickly draw up this fluid until the mixture reaches 
101. While this is being done the pipette should be twirled be- 
tween the fingers, thereby insuring a thorough mixing of the liquid. 
Stop suction and remove rubber tube. With a thumb closing one 



60 LABORATORY METHODS. 

end and the middle finger closing the other, shake well. The dilu- 
tion is complete, and the free end of the catheter may be pulled 
over the point, as suggested above. After reaching the office, 
another shaking is advisable, as the cells show a tendency to settle 
to the more dependent portions. Some persons prefer to remove 
the tubing entirely before leaving the home of the patient, and slip 
around the ends of the pipette a wide rubber band, which serves 
to prevent any leakage. 

Errors and Difficulties. — Solid columns of liquid are imperative. 
Air segments occur when the drop is too small or when the point 
of the pipette has not been sufficiently immersed. It may be impos- 
sible to draw any fluid into the tube, which indicates a plugged 
pipette, due either to an old clot or a piece of dirt, which may be 
seen by the use of a hand lens or with the naked eye. A little 
glacial acetic acid and a horse hair may establish a lumen, and, 
in case these fail, a fine hypodermic needle may be used, but care 
must be exercised that the point of the pipette is not broken off. 
The same methods may be employed when a column of the blood 
has coagulated in the tube. In case the initial blood column passes 
the proper marking, a towel touched quickly, but skillfully, to the 
point may take up the excess of blood and bring the top of the 
column down to the proper mark. 

Red Count. — After discarding four drops of the diluted blood, 
one small drop is blown into the clean counting chamber and the 
cover glass applied. The contact of the glass surfaces must be 
perfect, and this is shown by Newton's rings — a play of rainbow 
colors — seen best from an almost horizontal reflection of light. If 
the drop is too large, it spills out of the well and prevents the 
appearance of the rings. In such case the rings may be brought 
out by pressure on the cover glass, but will disappear if the pres- 
sure be removed. Permit the preparation to stand undisturbed 
for five minutes, so that the corpuscles may settle into the pens. 

There is practically no difference in the various rulings so far as 
the inner counting pens are concerned. Butler proposes that 
when the lines become faint they may be rendered very distinct by 
scraping off some graphite from a soft lead pencil, rubbing the 
powder over the surface of the disk, and then polishing it off with 
a soft handkerchief. 

The authors have tried with varying success at least half a dozen 
methods of counting red blood cells, but feel that still another 



VASCULAR DRAMAS. 



61 



method must be offered to the person who, although desiring the 
best results, must, for the sake of his patient, proceed with more 
haste than is usually required of many laboratory workers. The 
technic of this other method consists of a combination of several 
methods, and possesses the following advantages : 

1. Simplicity. 

2. By avoiding the square field methods, it obtains more nearly 
an average of the entire dilution. The most careful workers too 
often complain that the count in one area often varies exceedingly 
with other counts, and consequently many fields must be counted 
in order to find — what is, after all, the main result desired — an 
average. 

3. Prevents "losing the place." 



m 



tv& 



m 



M 



9 



m 



Fig. 15. — Scheme for rapid counting of red cells. 

Simple Method. Only four careful counts are made, and each 
of these includes ten squares, or forty in all. This may seem too 
few, but, if each of the four counts is made in a different portion 
of the preparation, there will be greater probability of striking an 
average than where several square areas are counted and the fact 
overlooked that, in spite of the best efforts, the cells lie more 
closely together in one area than in another. 

All corpuscles on the heavy lines and within the shaded areas are 
counted (Fig. 15). The beginning position is self-evident, and 
may be any one of a number of points, the new Turck ruling giving 
a choice of one hundred and forty-four such counts. This method 
guards against ' ' losing the place, ' ' a complaint that is old and well 



62 LABORATORY METHODS. 

founded. By using the proper ocular and drawing high the tube 
of the microscope, the field may be greatly magnified. Always use 
a low-power objective and narrow the diaphragm. In normal blood 
each of the little squares should average about 6.2 cells, which 
number is increased in polycythemia and decreased in anemia. An 
estimate of the number of red cells in each cubic millimeter of 
capillary blood should be made, and this computation, for diagnostic 
purposes, need not be accurate, but, while watching the progress 
of a blood disease, considerable care must be taken. Add the four 
countings, multiply by 2, and add four ciphers to the product. 
For example, suppose. the four counts w T ere respectively 60, 59, 57, 
and 60. The sum of these would be 236 ; multiplying by 2 gives a 
product of 472; adding the ciphers, we have 4,720,000 cells for 
each cubic millimeter. This should be done in a very few minutes, 
and, it is safe to say, no better average of the entire ruled disk 
could be taken. 

Cleaning Pipettes. — This should be done as soon as the count is 
finished, and the following method gives the best results : 

1. Attach rubber tubing to the pointed end of the pipette, re- 
moving it from the other end. 

2. Remove the remainder of the blood solution by blowing 
through the rubber tubing. 

3. Immerse the large end of the pipette in dilute acetic acid, 
suck the liquid into the pipette, and blow it out, repeating the 
procedure with absolute alcohol. 

4. Suck some ether into the pipette. 

5. Remove rubber tube and hold pipette firmly in hand. 

6. Because of the moisture ordinarily present in the exhaled air, 
ether can not be expelled by blowing, but by swift downward jerks 
of the hand. 

Only in case the glass mixing ball shows no tendency to adhere 
to the inner surface of the pipette, can the latter be placed away 
as clean. It may be necessary, if blood is still present, to repeat 
the entire technic, but, if only moisture interferes, the acetic acid 
wash may be omitted. Stronger acids, horse hairs, or even fine 
needles may be employed to remove the more obstinate clots. 

White Count. — Various bedside technics, where a hand lens is 
used, have been devised. They may serve some persons well, and 
should not be condemned, but the authors adhere to the use of the 
microscope for the following reasons : 



VASCULAR DRAMAS. 



63 



1. The older methods are simple, and white counting does not 
usually require haste, except possibly in infectious cases. 

2. More accurate results are obtained when privacy is secured 
and a microscope is used. Correct counts are not easily obtained 
in the environment of the sick-room. 

3. The microscope always distinguishes between cells and dirt, 
and also between red cells and white cells. 

It will, therefore, be seen that the rough methods of counting 
white cells are subject to serious limitations. The special white 




Fig. 16. — Spreading. 



After the blood has run along the edge of the spreader, the drop is 
pushed over the surface of the other slide. 



cell pipette and 1-percent acetic acid as a diluent are used, the 
technic being identical with that described for the red cells. The 
acetic acid destroys the hemoglobin, so that only the white cells 
may be seen, and brings out the nuclei of the white cells. Much 
yellow sediment indicates that the acid is not strong enough. It 
is often necessary to filter the acid solution at intervals of several 
months. 

Draw up the microscope tube to such a distance that the 
periphery of its field cuts exactly the corners of the large square 



64 



LABORATORY METHODS. 



millimeter, and permit the tube to remain in this position. In 
different portions of the preparation take five counts, including in 
each of these every white cell in the field. The sum of the five 
fields divided by 2, and with two ciphers added to this quotient 
gives the number of leukocytes in each cubic millimeter of blood. 1 
For example, suppose that the counts are 29, 35, 35, 32, and 35, 
their sum would be 166 ; this divided by 2 gives 83 ; adding two 
ciphers gives 8,300. This rapid method was proposed by F. J. 
Wright, of the Calumet and Hecla Hospital, Michigan. 

Making the Spread. — The most satisfactory method of prepar- 
ing blood films is that in which the slides, rather than cover glasses, 




Fig. 17. — Artifacts in blood films. A, pressure forms; B, elongations; C, poor fixation; 
D, dirt; E, normal stain; F, understating ; G, overstaining ; H, crenated cell; I, 
ghost or sh;dow. 

are used. The following technic takes the preparation to the stain- 
ing: 

1. Touch the edge of an end of a clean slide to a blood drop and 
transfer it to the face of another slide near its end. 

2. When the drop spreads along the edge of the smearer, push it 
over the face of the second slide (Fig. 16). 

3. Dry in air. 

4. Wrap in stiff paper and carry to the office for staining. If 
left exposed to flies, they will rapidly eat off the corpuscles. 

5. No fixation is necessary when using Wright's stain. 
Artifacts in Erythrocytes. — (Fig. 17.) Where fields are too 



1 A dilution of 1:40 is necessary. This proposition has reference to the large form 
of the white blood counter, which has a capacity of 21. In case, however, the small 
form of the white pipette is used, which has a capacity of 11, corrections must be 
made accordingly, but the former pipette is recommended as being more convenient. 



VASCULAR DRAMAS. 65 

crowded, certain irregular pressure forms may suggest poikilo 
cytosis. Parallel elongations of cells are due to pulling them out 
while making the smear. Bean-shaped forms are a direct result 
of poor fixation. A piece of dirt — usually black, rather than blue 
— may, by virtue of its position on an erythrocyte, suggest a normo- 
blast. Endoglobular degeneration is made evident by the tendency 
of certain erythrocytes to take the stain rather poorly. If, how- 
ever, all the red cells appear pale, there is sufficient reason to con- 
clude that the preparation has been understained. In case over- 
staining takes place, the delta may not be easily recognized. 

Staining the Blood Film. — The Wright's stain is simple, its use 
rapid, and its action all that can be desired. A little practice is, 
however, necessary, and the following technic will give the best 
results : 

1. Cover the film with a noted quantity of the staining fluid 
dropped quickly from a pipette or medicine dropper. 

2. After two minutes add to the staining fluid on the film the 
same quantity of distilled water with the medicine dropper, and 
allow the mixture to remain exactly two and one-half minutes. If 
the stain is all right, this should give an intense coloration to the 
cells. A longer period of staining invariably produces a precipi- 
tate. Eosinophilic granules are best brought out by a shorter 
period of staining. The quantity of the diluted liquid on the 
preparation should not be so large that some of it runs off. The 
water may show a tendency to gather in drops and roll off the 
stain, so that it may be necessary to mix it thoroughly with the 
latter before all is added. An ordinary medicine dropper full of 
the stain and the same amount of the water are usually sufficient. 

3. Pour off the liquid and wash the preparation in ordinary 
w T ater for thirty seconds, or at least until all precipitated stain is 
washed away and the preparation assumes a pink color. 

4. Dry between blotters. 

5. In several places, as may seem advisable, add cover glasses by 
means of balsam. If square or oblong cover slips are used, prac- 
tically all of the preparation may be covered. 

Authors' Slide Forceps. — Any one who has worked with the slide 
smear in preference to the cover glass preparation has been at a 
disadvantage in staining, as the Novy locking forceps, as well as the 
other varieties, can not be used with the ordinary slide. The 
authors have devised locking forceps which are sufficiently power- 



66 



LABORATORY METHODS. 



ful to render the drop staining method applicable to slide spreads. 
In the illustration (Fig. 18) a regular hemostatic forceps has been 
used, and the bends not only permit point coaptation, but guard 
against crushing the slide. 

Examination of Spreads. — This consists of a study of a stained 
smear under low and high power, especially under the latter. 
Proper diagnoses may be made by reference to the table on com- 
mon blood conditions (page 58). For a description of the stain- 
ing properties of the corpuscles see page 56. 





Fig. 18. — Authors' slide forceps. A, Pean's hemostatic forceps; B, shows alteration; X, 
prevents crushing of the slide when locking; Z, permits point coaptation. 



Differential Leukocyte Count. — Certain diagnostic and prog- 
nostic data may be obtained from the determination of the per- 
centage of the various types of white blood cells which may be 
present — an absolute increase in the polymorphonuclears being 
termed a leukocytosis, a decrease being termed a leukopenia, etc., 
each of the varied pictures representing some fact of clinical im- 
portance. 

For this work a mechanical stage is an aid, but not a necessity, 



VASCULAR DRAMAS. 67 

the latter statement being a direct contradiction of that usually 
made in text books. At least 300 white cells should be classified 
unless the probable condition is self-evident. The computation is 
easily made. For example, if 255 of these 300 cells were lympho- 
cytes, it follows that these constitute 85 percent of the entire white 
count — probably a lymphatic leukemia, etc. Whether a mechan- 
ical stage is or is not employed, the worker must never attempt to 
encroach upon a field whose cells have been previously numbered. 

Parasites and Microorganisms. — The presence of the Plasmo- 
dium is easily demonstrated by Wright's stain, which offers not 
only the most simple procedure, but brings out the most beautiful 
color contrasts. The body of the parasite stains blue, while the 
chromatin varies from lilac to red, or almost black. The chief 
source of error is mistaking blood platelets for endoglobular forms 
of the Plasmodium, as the former often lie upon or even within the 
red cell, but are of a homogeneous blue color. The plasmodium, 
when really seen, is rarely mistaken for anything else. It is not 
the purpose of this book to enter into a discussion of the morphol- 
ogy of this microorganism. 

To Prove Presence of Blood. — Take a stain. 

1. Place some of the stain on a glass slide. 

2. Add 1 drop of very dilute (.85-percent) salt solution. 

3. Evaporate very slowly at a low temperature. 

4. Add 4 drops of glacial acetic acid. 

5. Apply a clean cover glass. 

6. Again evaporate very slowly at a low temperature ; fluid 
should steam, but not boil. 

7. Add, twice, 3 drops of glacial acetic acid, and evaporate each 
time. 

8. Cool. 

9. Elevate cover glass and add 1 drop of glycerin. 
10. Examine. 

Blood, if present, is shown by the presence of dark-brown rhom- 
boid crystals, which may vary in size, and there will be a tendency 
of these crystals to rosette formation. This is called the hemin 
test. 

To Differentiate Fowl's from Mammal's Blood. — Take a blood 
clot. Tease out some of the clot in Marx 's fluid and examine under 
the microscope. Marx's fluid is made up as follows: 



68 LABORATORY METHODS. 

R Quinin hydrochlorate, 1 : 1,000 2 drams. 

Potassium hydrate solution, 33-percent 2 drams. 

Eosin, yellow , about 5 drops. 

Misce. 

Red cells should be stained pink. Those of the mammal appear 
as unnucleated circular disks, but those of birds and reptiles are 
oval and nucleated. 

To Prove Human Blood. — This can be done only by the pre- 
cipitin test, a difficult procedure, calling for expert assistance. 

Determination of Coagulation Time. — This may be roughly 
computed as follows : 

1. Allow several drops of fresh blood to fall on a clean glass 
slide. 

2. At intervals of a minute make tests by drawing a smooth 
white broom straw lightly through each drop. 

3. The coagulation time is reached when threads of fibrin tend 
to cling to the straw. The average coagulation time is about five 
minutes, and when it reaches or exceeds ten minutes it may be 
considered pathological, and operative procedures should be at- 
tempted with caution. In jaundice and hemophilia this time is 
greatly increased. The coagulation time may be experimentally 
or therapeutically shortened by the administration of gelatin or the 
calcium salts. 

Widal Test. — This is described in Diazo Versus Widal, page 104. 

Diagnosis of Carbon Monoxid Poisoning. — Treat the sample of 
blood with twice its volume of 1.3 specific gravity solution of caustic 
soda. Normal blood is changed to a dirty-brown, but, if carbon 
monoxid is present, a beautiful cherry-red will be seen. 

Tests Seldom or Never Attempted by the Practitioner. — 

1. Opsonic work. 

2. Wassermann reaction and its modifications. 

3. Detection of iodophilia. 

4. Searches for tropical microorganisms. 

5. Blood cultures; searching for microorganisms in the blood 
other than the plasmodium. 

6. Diabetes tests. 

7. Medico-legal work. 

Value and Limitation of These Tests. — A correct interpretation 
of these tests, with a more or less complete application of the va- 
rious procedures, will prove of inestimable value in many diagnos- 



VASCULAR DRAMAS. 69 

tic and prognostic difficulties. A "blood analysis" should not, 
however, be overestimated, as a blood picture may be ever so beau- 
tiful and characteristic, teeming with symptomatic therapeutic 
indications, and notwithstanding this appearance fail to indicate 
the chief etiological factor. It is often only by a series of exam- 
inations that the case in question may receive a rational, scientific 
treatment. 



70 



LABORATORY METHODS. 




CHAPTER V. 



CHEMISTRY AND BIOLOGY OF THE GASTRIC JUICE. 



Apparatus. — Analysis of the gastric contents shows but few 
thing's of value to the clinician or the general practitioner, but 
these are very important in making a correct diagnosis of a stom- 
ach disease. In order to make these tests w r ith ease and accuracy, 
only a few special pieces of apparatus and a few reagents are nec- 
essary : 



1. Buret, graduated to 50 cc, with 

stop-co.ek or pinch -cock. 

2. Sodium hydrate, VI n normal. In 

order to obtain dependable re- 
sults, this solution must be ac- 
curate. Prepared quantitative 
solutions are obtainable from 
reputable chemical firms, or may 
be prepared by an expert phar- 
macist. 

3. Dimethylamidoazobenzol, a weak 

aqueous solution. 



4. Phenolphthalein, a 1 -percent al- 

coholic solution. 

5. Gunzburg's reagent. 



6. 



Ferric chlorid. 

Stomach tube with bulb. 

Evaporating dish. 

Delicately graduated pipette, 10 

cc. 
Slides and cover glasses. 
Microscope and accessories. 



Contraindications for Stomach Washing. — 

1. Recent hemorrhage from the stomach. 

2. Diseased esophagus, which may lead to perforation with the 
tube. 

3. Aortic aneurism. 

4. ]\Iarked arteriosclerosis. 

5. Angina pectoris. 

6. Other examinations may precede the passing of a stomach 
tube. Tabes dorsalis, tuberculosis, nephritis, and -anemia may 
produce grave gastric symptoms. A detached tenth rib may point 
to a gastroptosis, or certain stigmata and symptoms may indicate 
the presence of hysteria. 

Preparation. — At the regular breakfast time for the patient he 
should be given an Ewald test meal consisting of two slices of white 

71 



72 LABORATORY METHODS. 

bread, with the crust removed, without butter, and a glass of warm 
water or weak tea. The bread, which should be somewhat dry, 
must be well chewed, and not dipped into the liquid, but washed 
down with it. The contents are to be removed forty-five minutes 
after the meal. 

Passing the Tube. — It is advisable to inform the patient that the 
operation will be unpleasant, but not painful. The tube should be 
warm, and should be lubricated with a very thin coat of glycerin. 
The patient should be seated in a chair with arms, so that he can 
grasp them with his hands, his head thrown well back, and his 
mouth wide open. Stand on the right side of the patient, with 
your left arm around his head, so that the left hand can be used 
to guide the tube as it is passed. Seize the tube about five inches 
from the tip, and place the tip, pointing downward, against the 
posterior pharyngeal wall, at the same time directing the patient 
to swallow. Then, holding the tube with the fingers of the left 
hand to keep it from being coughed out, push it steadily and rather 
rapidly down the esophagus. At the cardia it will meet with a 
little resistance, but this is soon overcome, and the tube will enter 
the stomach. If it meets a firm resistance, it is most probably 
against the stomach wall, and should be drawn back an inch or 
two. Direct the patient to close his lips and breathe rapidly, 
which will distract his attention and help to prevent retching. If 
the patient is very irritable or nervous, spray the pharynx with a 
weak cocain solution a few minutes before passing the tube. Gen- 
erally, if care is taken and the operator does not use too much 
haste, no trouble will be found in passing the tube. 

To Remove the Contents of the Stomach. — Grasp the tube with 
the right hand (Fig. 20), closing the tube at A with the thumb and 
forefinger of the same hand. Compress the bulb and close the tube 
at B with the fingers of the left hand, and immediately release at 
A. The bulb will expand, and the contents will be drawn through 
the tube into the bulb. Repeat this process until no more fluid is 
obtained. 

If, after removing the contents for analysis, it is desirable to 
wash out the stomach for therapeutic purposes, the free end of the 
tube can be placed in a vessel of warm water, and the valve action 
of the fingers reversed so as to pump water into the stomach. In 
the same way the tube can be used as an air compressor to dilate 
the stomach, so that it can be outlined by percussion. 



CHEMISTRY AND BIOLOGY OF GASTRIC JUICE. 



73 



The tube is removed by a slow, steady pull. Give the patient a 
drink at once and food in about half an hour. 

Examination of the Contents. — The material obtained should be 
examined as soon as possible after removal in order to avoid the 
artifacts of continued fermentation. Note the odor at once to 
determine if there has been any putrefaction. Filter the material 
through a single sheet of filter paper. Use a large funnel, as the 
filtrate will pass through slowly at best. Reserve some of the Tin- 
filtered liquid for microscopic examination. 

Macroscopic Examination. — Quantity. This is a variable fac- 
tor, and therefore no standard of normal can be given. It will 
vary with the motor activity of the stomach and with the amount 




Fig. 20. — Method of removing contents from stomach. 



of secretion. A large amount may signify retention or hyperse- 
cretion, or both, and a small amount may indicate hypermotility or 
scanty secretion. The chemical and microscopical examinations 
must be made as a check for the quantity of material. 

Color. The. normal color after an Ewald meal is yellowish- 
white. The presence of red blood indicates hemorrhage at the time 
of passing the tube, and may or may not have been caused by the 
tube, but is generally considered as a sign of congestion. Dart 
brown and black specks or particles indicate hemorrhages previous 
to the passing of the tube, and are partially digested blood clots. 
A greenish-yellow tinge is caused by bile, which is often forced 



74 



LABORATORY METHODS. 



back into the stomach by the straining and retching when the tube 
is passed. 

Physical Characteristics. The material in health consists 
chiefly of finely divided particles of the food taken. Mucus, of 
which there should be only a small amount, is recognized by its 
tenacious character. Food taken at a previous meal, or even a day 
or two before, can often be identified by bits of vegetable or meat 
fibers, seeds, and skins of fruits or vegetables. Of these the most 
easily recognized are the red skin and seeds of tomatoes and the 
skins of raisins. 

Pieces of mucosa or tumor are difficult to recognize, and suspi- 
cious materials should be sectioned and examined histologically. 
(See Essence of Tissue Diagnosis, page 78.) 




® 



G 



© 



© 





t 



V) 



N 



Fig. 21. — Microscopic elements of major and minor import in stomach analyses. Normal 
findings — A, muscle fibers from food: B, vegetable cells from food; C, starch grains 
of food; D, fatty needles from food; E, squamous epithelium from mucosa. Patholog- 
ical elements — F, white blood cells, normal in small numbers; G, red blood cells, 
rarely normal even in very small numbers; H, yeast cells; I, sarcines; J. Oppler 
Boas bacilli, longer rods. 



Microscopic Examination. — Take some of the material which 
remains in the filter, and press a small amount of it between two 
large glass slides or between two plates of clear glass. Examine it 
first with low power for connective tissue fibers and pieces of mu- 
cosa and tumors. Examine with high power for the following 
(Fig. 21) : 

Blood Cells. These may be in their normal shape or crenated- 



CHEMISTRY AND BIOLOGY OF GASTRIC JUICE. 75 

Yeasts. These are elongated bodies, three or ten microns in 
length, with similar smaller processes budding out from them. 

Sarcines. Sarcines are rather large cocci, which form "cotton 
bale'' clusters as they divide. 

Oppler-Boas Bacilli. These are long, heavy bacilli, which often 
occur in pairs, and may be found in long chains or forming angles 
with each other. 

Chemical Analysis. — Qualitative. Free hydrochloric acid. 
Mix 3 or 4 drops of Gunzburg's reagent and an equal amount of 
the filtrate on a porcelain shell or on the bottom of a clean evapo- 
rating dish, and heat slowly over a flame, holding the dish with 
the fingers in order not to burn the materials. A deposit of minute 
red crystals at the edge cf the mixture gives a distinct rose-red 
color when free hydrochloric acid is present. This is a certain 
reaction. 

Lactic acid. In a large test tube mix 5 cc. of the nitrate with 
50 cc. of distilled water, and add 2 drops of ferric chlorid (5-per- 
cent aqueous solution). If lactic acid is present, a greenish-yel- 
low color appears. 

These two qualitative tests are the only ones of any value to the 
practitioner. 

Quantitative. The most important quantitative analyses are 
for free hydrochloric acid and total acidity, and these may be made 
together. To 10 cc. of the nitrate in a beaker or whisky glass add 
1 drop of dimethylamidoazobenzol indicator. Starting with the 
buret rilled with the N 10 NaOH to the cc. mark, add the NaOH 
to the filtrate, a drop at a time, stirring constantly. When the 
pink color has disappeared, all the free HC1 has been neutralized, 
and the reading of the buret gives the number of cubic centimeters 
cf NaOH used in the neutralizing. As only .10 cc. of filtrate were 
used, this number multiplied by 10 will give the percentage of 
HC1 in 100 cc. of the filtrate in terms of N/10 NaOH. For ex- 
ample, if 4.5 cc. N/10 NaOH are required to neutralize the free 
HC1 in 10 cc. of the stomach washings, then 10X4.5, or 45, is the 
percentage of free HC1 in terms of N/10 NaOH. 

To the same filtrate now add 2 drops of phenolphthalein, and 
again titrate with NaOH until the first permanent pink occurs. 
The reading of the buret X 10 now indicates the total acidity, and 
this, minus the free acid, is practically the amount of combined 
acid. 



76 LABORATORY METHODS. 

In case that enough filtrate can not be obtained to furnish 10 cc. 
for the quantitative tests, use as much as possible and adjust the 
figures according to the following example : 

Number of cubic centimeters used : 10 : : NaOH used : x, x equal- 
ing the number of cubic centimeters of NaOH that would be re- 
quired by 10 cc. of the filtrate. 

If 6 cc. of the nitrate required 3 cc. NaOH for free HC1, then 

6 : 10 :: 3 : x 

x = 5 
5 X 10 = 50= percentage of free HC1. 

If 2 cc. more of NaOH are required for the total acidity, then 

6 : 10 : : 5 : x 
x = 8.33 
8.33 X 10 == 83.3 = percentage of total acidity. 
83.3 — 50 = 33.3 = percentage of combined acid. 

Interpretation of Findings. — Yeasts require time to grow, and 
may be considered as indicative of some retention and fermenta- 
tion. 

Sarcines can grow in free acid, and are generally indicative of 
hyperacidity. 

Oppler-Boas bacilli are generally found in hypoacidity, as they 
do not grow well in free acid. They produce lactic acid, and 
where one is found the other may generally be expected. 

In ulcer of the stomach the free and combined hydrochloric acids 
are increased, hemorrhage is common, and sarcines may be pres- 
ent. There is often a hypersecretion, so that large amounts of 
material may be obtained in washing. Eetention will be shown. 
No lactic acid is found in the majority of cases. 

Carcinoma of the stomach is accompanied by a diminished hy- 
drochloric acid content, and often no acid is found. The Oppler- 
Boas bacillus is nearly always found, and is considered by some as 
pathognomonic of carcinoma. The secretion is scanty, and only 
small amounts of material are obtained after a test meal. Lactic 
acid is generally found. 

Chronic gastritis gives a variety of findings, and the most con- 
stant of these is mucus in large amounts after a meal or after a 
fast of a few hours. 



CHEMISTRY AND BIOLOGY OF GASTRIC JUICE. 77 

Gastric neuroses may be indicated by the presence of abnormally 
la roe or deficient quantities of stomach acids without the presence 
of pathological elements. 

Sources of. Error. — The main sources of error may be traced to 
inaccurate quantitative test solutions. When making; the iron test 
for lactic acid it is advisable to run a control test, as the final green 
color is often difficult to determine. It may be very tedious to 
filter the sample, which is especially the case when much mucus 
is present, and it is suggested to pass it through a single piece of 
cheese cloth before using the paper. 

Value and Limitations. — A thorough physical examination 
should precede every stomach analysis. If this fails to reveal the 
nature of the disorder, and there are no contraindications, the physi- 
cian should proceed with the analysis of the gastric juice and ex- 
pect to learn much from the examination. 

It must be borne in mind that the results of a laboratory exam- 
ination should not overshadow those obtained by other measures, 
and that each link in the chain of evidence must be equally strong 
in order that a safe diagnosis may be established. 

Less Frequently Applied Procedures. — The stomach whistle, the 
gastric bucket, and mirror have yet to earn a place in the equip- 
ment of the general practitioner. 

Many chemical and microscopic examinations of scientific inter- 
est have been omitted in order that the more useful procedures in 
diagnostics might be emphasized. 



Note — Weinstein (Archiv. of Diag., July, 1912) calls attention to the fact that many- 
valuable points may be gained by the gross appearance of the stomach contents. Thus, 
he states, chronic gastritis is characterized by thick contents with considerable amounts 
of stringy, glassy mucus. In the hyperacid states, the prrticles of bread are heavy; and 
fall to the bottom of the vessel like grains of sand. In hypoacidity, the particles of 
bread are more likely to be flaky and float. 



CHAPTER VI. 

ESSENCE OF TISSUE DIAGNOSIS. 

The subject matter of this chapter has, for convenience, been 
treated under two divisions — "Essence of Frozen Sections" and 
"Essence of Celloidin Sections." The first division treats of the 
rapid tissue diagnosis in the operating room by means of frozen 
sections, but, as it is often desirable to proceed more slowly, the 
treatment in the second division offers a simple technic. Both 
methods have been used by the authors with the most gratifying 
results. Curettings from the cervix uteri, specimens from sus- 
pected breast tumors, and other tissues are examined by the first 
method and a diagnosis made within five minutes, provided that 
the technic has been thoroughly mastered by practice with pieces 
of steak, lumps of sausage, etc. 

In this chapter no attempt is made to teach pathology, but the 
practitioner may, however, gain much information on this subject 
by frequent reference to the high-class atlas. 

ESSENCE OF FROZEN SECTIONS. 

Apparatus. — The articles forming the equipment have been listed 
in the order of their use. 



1. Whisky glass. 

2. Formalin, 10-percent. 

3. Scalpel. 

4. Forceps. 

5. Microtome. 

6. Common bottle corks. 

7. Gum arabic (concentrated aqueous 

solution) and pipette. 

8. Tube of ethyl chlorid. 

9. Section knife, or razor. 



10. Watch glass, or saucer. 

11. Distilled water. 

12. Needle and section lifter, or sub- 

stitutes. 

13. Slides. 

14. Thionin stain and pipette. 

15. Cover glass. 

16. Filter paper. 

17. Microscope and accessories. 



References. — Mallory and Wright : Pathological Technique ; Diirck : Atlas of Patho- 
logic Histology; Warthin : Laboratory Work in Pathology; Hall and Herxheimer: 
Pathological Methods. 

78 



ESSENCE OF TISSUE DIAGNOSIS. 



79 



With the exception of the microscope and. small microtome, the 
entire outfit may be kept in a small box about 12x5x3 inches, which 
may be easily taken to the hospital or home of the patient. "While 
quite as reliable results may be obtained with this outfit as with the 
more expensive apparatus, it must not be inferred that the speci- 
mens will be beautiful. The frozen section is, at best, thicker than 
the celloidin cutting, but, so far as diagnosis is concerned, shows 
quite as much. 

Microtome.— A perfected microtome is a valuable, but expensive, 
instrument. The authors have tried many substitutes, and have 
found that a thin section, made with a razor, using a pair of tissue 
forceps and an ethyl chlorid spray, is satisfactory. Herman sug- 
gests the use of a small rubber band wrapped around the points of 






Fig. 22. — Substitutes for the perfected microtome. A, Relihan's suggestion, costs 10 
cents; B, hand microtome, costs $6; C, table microtome, costs $12 to $14. 

the forceps (Fig. 22), the upper edges of the blades serving as ways 
and the razor being thus supported at a cutting angle. He says: 
"The first section will be too thick, but, after freezing the third 
or fourth time and repeating the same procedure, I have had no 



80 LABORATORY METHODS. 

trouble in getting thin sections. The contraction of the forceps 
blades seems to be so much quicker than the tissue contraction that 
the projecting portion is about the right thickness for a section. 
By going through this procedure . . . these sections answered 
my purpose perfectly." 1 The hand microtome (Fig. 22) answers 
for thick sections. 

The authors recommend the use, whenever possible, of a table 
microtome, which is supplied by all optical firms, and with it prac- 
tically every section is thin enough for study. Although the 
description is confined to this instrument, all directions may be so 
modified as to be applicable to any substitute. 

Freezer. — Certain ether and rhigolene freezers, ranging in price 
from $6 to $10, are on the market and give good results. They 
do not, however, freeze under certain atmospheric conditions, nor 
are they easily manipulated when haste is desired. The ethyl 
chlorid spray, used in minor surgery, is more convenient, can be 
had at a modest price, and the results of its use are so encouraging 
that it is recommended to the general practitioner. 

Knife. — Any razor will answer, especially if one side of the 
blade is flat — not hollow ground. The authors have successfully 
used the new Bausch & Lomb chisel knife, an inexpensive instru- 
ment, for frozen sections. 

Fixing and Mounting. — With the scalpel and tissue forceps the 
specimen is cut into a small block about the size of a pea and 
dropped immediately into a tumbler containing 10-percent forma- 
lin (4-percent formaldehyd), when it is carried to where the ex- 
amination is to be made, and the short time that it is in the solu- 
tion will aid in freezing it. Alcohol delays freezing and does not 
answer the purpose. The tissue is then seized with the forceps 
and dipped into a concentrated aqueous solution of acacia, after 
which it is placed on the top of an ordinary cork previously 
mounted on the microtome. 

Freezing and Sectioning. — The tissue, in its gummy capsule, is 
frozen at once by using the ethyl chlorid spray. Very little of 
this liquid is necessary if the operator blows vigorously on the 
tissue, and under such treatment it should harden almost in- 
stantly. Sections are quickly obtained, and should be as thin as 
possible. 

Staining. — The sections are dropped immediately into some dis- 



1 Journal of American Medical Association, November 6, 1909. 



ESSENCE OF TISSUE DIAGNOSIS. 81 

tilled water, which should not be too cold if good stains are de- 
sired. Place a section on a clean slide and add immediately several 
drops of carbol-thionin. Permit staining to proceed for at least 
one minute, and take up excess of fluid with a little filter paper. 
Add a clean cover glass and examine under low power. 

Preparing the Stain. — The stain is so easily made up that the 
powdered thionin is recommended. The making of the liquid 
stain, as described by Strouse, is as follows : 

1. Take 2 grams of pure thionin. 

2. Make up a saturated aqueous solution. 

3. Allow the solution to stand at least six hours, stirring occa- 
sionally. 

4. Filter. 

5. Mix the clear solution with equal parts of 2-percent phenol. 
The stain improves with age, but an occasional filtering may be 

necessary. 

Technic— A brief outline of the technic of quick tissue diagnosis 
is as follows : 

1. Cut into small block. 

2. Transfer in 10-percent formalin. 

3. Mount on cork in acacia solution. 

4. Freeze with ethyl chlorid. 

5. Section. 

6. Float in warm distilled water. 

7. Drop on slide. 

8. Add stain. 

9. Add cover glass. 
10. Examine. 

Examination of Section. — Too much must not be expected from 
this method, and even a person with some experience in the ordi- 
nary technic may be confused at first. Compared with celloidin 
sections, these are much thicker and do not stain so well. With a 
little patience, however, and when applied to certain selected cases, 
it proves a very valuable addition to the medical laboratory. 
Examinations should • be made principally with a low-power ob- 
jective, especially in diagnosticating tumors. The thionin stain is 
usually not intense, so that it may be advantageous to narrow the 
diaphragm. Nuclei are stained a dark-blue color, while the proto- 
plasm takes on a reddish purple tinge. 

Errors in Technic. — Alcohol can not be substituted for formalin, 



82 



LABORATORY METHODS. 



as it prevents or delays freezing. The refrigeration must be com- 
plete, and the tissue should, figuratively speaking, become as hard 
as a rock before any attempt at sectioning is undertaken. At the 
first indication of thawing, the tissue should be again frozen if more 
sections are desired. A dull knife and a straight cut cause the 
tissue to crumble. The knife should be drawn at an angle best 
estimated by experience. Staining should cause no trouble, and a 
thioniu solution which refuses to stain after it has been prepared 
two weeks should be rejected. Neither the stain nor the wash 
which precedes it should be cold if good results are desired. 

Other Freezing Methods. — In hospitals and other institutions, 
where expense must serve convenience, the carbon dioxid freezing 
tank and attachment are used. 

Value and Limitations. — The freezing methods are used at pres- 
ent only in these diagnostic indications which have been previously 
considered (page 78), and are not advised where haste is not 
necessary, as the sections are thick, stain poorly, and are often of 
no service in the differentiation of cellular elements. In its 
sphere, however, the value of the rapid method can hardly be over- 
estimated. The technic can not be mastered in a few minutes. 
Repeated attempts with controls — tissues of known sources — 
should be made, and the apparatus collected and held in reserve 
for the time of need. 



ESSENCE OF CELLOIDIN SECTIONS. 



Apparatus. — 

1. Absolute alcohol. 

2. Blotter system. 

3. Carbol-xylol. 

4. Celloidin shavings. 

5. Corks. 

6. Distilled water. 

7. Eosin (1-percent solution). 

8. Ether. 



9. Glass covers. 

10. Mayer's hemalum solution. 

11. Microscope and accessories. 

12. Microtome and knife, or substi- 

tutes. 

13. Section lifter. 

14. Staining dishes, or substitutes. 

15. Whisky glasses. 



Of all the celloidin methods, this seems to be the most simple 
and rapid, as well as sufficiently accurate, technic for those desir- 
ing to do this kind of work. 

Preparing Celloidin Solutions.— Shering's celloidin shavings 
are sold in 1-ounce bottles by optical firms. A stock solution is 
prepared by dissolving about 2 drams of the shavings in a 6-ounce 



ESSENCE OF TISSUE DIAGNOSIS. 83 

bottle filled with equal parts of absolute alcohol and ether. Keep 
well stoppered, shaking occasionally, and a perfect solution should 
occur within twenty-four hours. This is the stock solution, or the 
thick celloidin solution. To prepare the thin celloidin solution, 
dilute some of the stock solution with an equal quantity of a solu- 
tion of equal parts of absolute alcohol and ether. 

All of these solutions are very volatile and highly inflammable, 
and must be kept in well-stoppered bottles. When used in glasses 
or staining dishes, the vessels should be covered with small squares 
of window glass, similar to those used Avhen examining urine sedi- 
ments. In case the solution must stand over night, a little petro- 
latum should be placed around the upper edge of the vessel before 
applying the cover. 

Rapid Hardening and Infiltration. — This method, somewhat 
modified, is used in the pathological laboratory at the University of 
Michigan. Each solution may be kept in a small tumbler, but 
evaporation must be avoided. A piece of the tissue about the size 
of a pea is selected and passed through the solutions as follows : 

1. Three changes of absolute alcohol during an hour. 

2. Equal parts of ether and absolute alcohol, one-half hour. 

3. Thin celloidin over night. 

Imbedding. — Remove the tissue from the thin celloidin and place 
it on the flat surface of a wide, but not too thick, cork — a cork 
similar to those used in small salt-mouthed bottles. Permit the 
celloidin to partially "set," thus gluing the preparation to the 
cork, and then pour on it a little thin celloidin and blow on it. By 
repeating this process several times the celloidin may be built up 
around the tissue. Now "float the cork, tissue face downward, in 
absolute alcohol for five hours or longer. 

Sectioning. — (See page 80.) Do not use the chisel knife, but 
an ordinary microtome knife or a razor. In case the latter is em- 
ployed, at least the under surface should be flat, and the upper 
surface must be kept covered with alcohol during the cutting. The 
section should be made by one continuous cut, using a good "cut- 
ting angle." Sections' are placed in ordinary alcohol before stain- 
ing. 

Systematic Staining (Authors' Method). — By using picrocar- 
min, less time will be taken, but the results are not nearly so 
good as where hemalum and eosin are used. Instead of staining 
dishes, watch glasses or saucers may be used. Small tin ointment 



84 LABORATORY METHODS. 

boxes possess many advantages, and, if they are new, the sections 
are easily seen against the bright tin background. Before begin- 
ning any portion of the staining, each dish should be supplied with 
its solution, covered with a glass plate, and be arranged in the 
order of their use. A watch or clock should be constantly in full 
view. The plan of systematizing here given has many advantages 
over the more haphazard methods. 

Each dish should rest on a small piece of blotter, which not only 
serves to keep the laboratory table clean, but on it may be placed 
the number of the dish, the name of the solution, and the staining 
time. This arrangement will not only keep the worker from be- 
coming confused, but, if laid aside between analyses, it will not 
always be necessary to read up the method when each tissue is 
examined, which is of importance to the busy practitioner. The 
following procedure not only illustrates what is meant by the 
"systematic" method, but serves to describe the routine of celloidin 
staining as recommended to the general practitioner. 

1. Hemalum, one to ten minutes — nuclear stain. 

2. Distilled water, one minute — wash. 

3. Eosin, one to five minutes — cytoplasm stain. 

4. Distilled water, one minute — wash. 

5. Ninety-five-percent alcohol, one minute — dehydrating. 

6. Absolute alcohol, one minute — dehydrating. 

7. Carbol-xylol, one minute — clearing. 

8. Mount in balsam. 

The duration of staining depends on the strength of stain and 
kind of tissues. With a little practice this method should give no 
trouble. The nuclei should appear as dark-purple spots on a pink 
cytoplasm. In case it is not desired to keep the specimens, the 
technic may be stopped short of the alcohols and the examination 
be made in water. 

Curettings. — These may be prepared for examination by either 
the freezing or celloidin method, several pieces, instead of one, be- 
ing mounted in acacia or .imbedded in celloidin respectively. 

Paraffin Imbedding. — A description of this method is left to the 
larger text books. It is true that very beautiful specimens may be 
obtained by the use of this substance, but for a small number of 
specimens it can hardly be recommended. Its application neces- 
sitates the purchase of expensive equipment, and it is a slow method 
so far as diagnosis is concerned. 



CHAPTER VII. 



DETECTION OF THE COMMON POISONS. 



Apparatus. — 

1. Ring stand. 

2. Evaporating dish, or porous porce- 

lain plate. 

3. Marsh apparatus. 

4. Test tubes, stand, and holder. 

5. Funnel and filter paper. 

6. Reagents, which should be chem- 

ically pure and obtained in small 
amounts from reliable companies 
— carbolic acid, chromic acid, 
hydrochloric acid (arsenic free), 



nitric acid, sulphuric acid, am- 
monium hydroxid, calcium chlo- 
rid, powdered charcoal (wood), 
ferric chlorid, Nessler's reagent, 
phenolphthalein (1-percent alco- 
holic solution), potassium bi- 
chromate (dichromate) , sodium 
hypochlorite (should be made up 
fresh by pharmacist), stannous 
chlorid, sweet oil, zinc (arsenic 
free). 



In this book are described only the good tests, which are usually 
the simple ones, and, with a view of limiting expense, the selections 
of tests have been made in such manner that one reagent may be 
available for the detection of several different drugs. For exam- 
ple, the ferric chlorid can be used in the identification of morphin, 
opium, and carbolic acid, and the same reagent may be used in 
urinalysis. As far as possible the authors have avoided indicating 
the rare and more costly reagents. 

These are sample tests, and are made for the purpose of satisfy- 
ing the physician or his patient. Only an isolation of the poison 
in its pure form is accepted as expert testimony, and it is not the 
object of this book to enter that field. 

There are certain limitations to the value of many of these tests, 
but these are not always pointed out because substances giving 
similar reactions are not usually met under the same conditions. 
For example, the physician observes certain symptoms in a patient 
which he believes to be due to phenol poisoning, and at once begins 
examining samples of the drug, food, or beverage under suspicion. 
The fact that anilin might give a similar reaction seems hardly to 



References. — Peterson and Haines: Legal Medicine and Toxicology; Edmunds and 
Cushny: Experimental Pharmacology; Autenrieth: Detection of Poisons; Riley: Toxi- 
cology; Tanner: Poisons. 

85 



86 



LABORATORY METHODS. 




DETECTION OF THE COMMON POISONS. 87 

enter into the question. So far as the common poisons are con- 
cerned, each reaction is specific, and should at once put the physi- 
cian on the right track, but such conclusion would not be accepted 
in a court as expert evidence. While these reagents are few and 
not expensive, they are not always to be obtained by the physician 
at a moment's notice, and, if he has a considerable amount of this 
work to do, he should keep on hand a sufficient quantity to meet his 
needs. 

All poisons are not freely soluble, so that the sample should not 
be filtered. Much better results may be obtained by straining out 
the larger pieces of foreign material, and decantation may often 
serve this purpose. Only the most common poisons are included, 
and it has been deemed advisable to omit aconite and other poisons 
of perhaps secondary import for the reason that they are not 
usually employed to destroy human life. For the sake of sim- 
plicity, names of the tests are not given, as many have been modi- 
fied to such an extent that it would be difficult to give a definite 
nomenclature. 

In the poison tests distilled water should, if possible, be used, 
although it is not absolutely necessary. In case ordinary water is 
used, it should contain neither lead nor any other common poisons 
— in other words, it should be gocd drinking water. 

Unfortunately there are no simple tests for wood alcohol and 
antipyrin which can be properly recommended, while several other 
tests — for example, the ergot test — are none too satisfactory. 

Sources of Every-Day Poisons. — "Dopes" are neither always 
served "straight," nor are they invariably so preferred by suicides. 
Some of the mere common mixtures or adulterations are : 

Abortifacients — ergot, yellow phosphorus. 

Bad whisky ("rot gut") — laudanum, wood alcohol. 

Bitters (bracers) — strychnin, alcohol in large proportion. 

Bug poisons — arsenic, mercury salts. 

Coke, coc, certain colas, fatigue powders — cocain. 

Cough syrups, soothing syrups, papine — morphin. 

Drug cures — morphin, cocain, alcohol, hyoscin, etc. 

Eye drops, clap medicines — zinc salts, silver salts. 

Headache powders — acetanilid, antipyrin. 

Knock-out drops — chloral, laudanum. 

Liniments — certain ones have been found to contain wood alco- 
hol. 



08 LABORATORY METHODS. 

Lye — fixed alkalies. 

Matches — yellow phosphorus (some of the safety matches con- 
tain the nonpoisonous reel phosphorus). 

Paris green — a mixture of copper acetate and copper arsenite. 

Rat poisons — arsenic ; some contain phosphorus, others contain 
cultures of the Danycz virus or certain strains of paratyphoid 
bacilli. 

Sex stimulants — cantharidin. 

Skin ' ' beautifiers ' ' — arsenic. 

Inasmuch as many poisons have been omitted, there has been 
no attempt at classification other than an alphabetical arrange- 
ment. For convenience, the names of necessary reagents precede 
the test. The letter "x" refers to the unknown substance. 

It is not necessary that percentage solutions should cause con- 
fusion. In qualitative tests, only approximate values are intended 
unless otherwise stated. A practical method of estimating these 
solutions is given in General Information, page 192. 

Acetanilid. — Concentrated hydrochloric acid, concentrated aque- 
ous solution of carbolic acid, ammonium hydroxid solution, very 
dilute chromic acid solution. Boil "x" with about y 2 dram of 
concentrated hydrochloric acid and cool. Add 1 dram of the car- 
bolic solution, and then add a few drops of the chromic solution. 
After two minutes a dirty purple or red should appear. Add a 
few drops of the ammonium hydroxid solution and shake. Within 
two minutes a greenish or indigo color should appear. 

Alkalies Versus Mineral Acids. — Phenolphthalein (1-percent al- 
coholic solution). 

1. Alkalies have a soapy "feel." 

2. Nitric and hydrochloric acids have characteristic odors. 

3. Acids act on carbonates with gas formation. 

4. Acids show a tendency to exhibit a sour taste even in very 
dilute solution. 

5. Test with phenolphthalein. Alkalies in very small amounts 
impart to this indicator a beautiful red color, whereas acids produce 
no effects. 

Alcohol. — Sulphuric acid (50-percent), potassium bichromate. 
Add to % dram of "x" % dram of the acid, and drop into the 
mixture a crystal of the bichromate. A green color should appear. 

Ammonia. — Hydrochloric acid. Dip a stirring rod into the acid 
and hold near "x. " White fumes should appear. The charac- 



DETECTION OF THE COMMON POISONS. 



89 



teristic odor of ammonia as well as the gas is lost on standing. 
Samples for examination should be tightly corked. 

Arsenic. — Hydrochloric acid, zinc, sodium hypochlorite, calcium 
chlorid. Mix "x," 4 drams of hydrochloric acid, several pieces of 
zinc, and 1 ounce of water (distilled or drinking) in a small Marsh 
apparatus, and cork. Do not inhale fumes. Do not light gas 
until you are sure all air has been expelled from the bottle. Test 
by collecting samples in inverted bottle, and, when these will burn 
without explosive violence, all air has been removed. 

Fig. 24 shows how a Marsh apparatus may be constructed with 
a bottle, a cork, and a graduated 5-cc. pipette, 
and illustrates how the hydrogen must be collected 
for testing. There must be no leaks in the ap- 
paratus. Now carefully light the gas, as, in case 
any air is present, a violent explosion will occur. 
Be careful to inhale none of the fumes, which 
give off a garlic-like odor and are very poisonous. 
Hold one of the unglazed porcelain plates in the 
flame, and, if a sublimate is deposited which is 





Fig. 24. — Improvised Marsh apparatus. A, bottle used as 
generator; B, pipette, such as is used in stomach tests, 
the expanded portion of which contains calcium chlorid 
crystals; C, bottle which serves as a receiver for col- 
lecting and testing the hydrogen. 

soluble in sodium hypochlorite, arsenic is present. 
Some difficulty may be experienced in intro- 
ducing calcium chlorid into the enlarged por- 
tion of the tube. Small crystals should be used, 
and it is not necessary, for practical purposes, 
that all these lie neatly in the enlarged portion of 
the tube, nor is it imperative that this enlargement should be en- 
tirely filled. If a thistle tube can be obtained and passed through 
the cork, its lower end- diving well below the liquid, fresh acid may 
be added from time to time as necessary. The amounts of reagents 
need not be accurate, and necessarily vary with the size of the bot- 
tle. \Yhen cleaning apparatus, avoid inhaling odors. Throw 
away the zinc. 




90 LABORATORY METHODS. 

The following instructions should be observed. 

1. Have all connections tight. 

2. Do not inhale fumes. 

3. Do not explode your apparatus. 

4. When lighting the gas, use a long rod to support match in 
order to guard against injury if an explosion should occur. 

Atropin, Hyoscin, Etc. — Both dilate the pupil of a dog's eye. 
When a sample of atropin is heated, a honey-like odor is given off, 
which may be intensified by the addition of oxidizing agents. 

Bacteria. — It seems that the future toxicologist must be trained 
to recognize certain pathogenic bacteria, especially the typhoid ba- 
cillus. Their isolation and identification are tedious, discourag- 
ing, and often impossible, even for the expert. The physician 
should bear in mind this new method of voluntary contamination 
of food and prescriptions. 

Cantharidin. — Sweet oil. Dissolve some of the suspected ma- 
terial in hot sweet oil, and shake well. When the oil rises to the 
top of the mixture, decant it into a separate vessel. Apply sev- 
eral drops to some adhesive plaster and place it on the chest, allow- 
ing it to remain in position about ten hours. A positive test is 
shown by the presence of a blister, or at least a severe reddening, 
where each drop touches the integument. Alkalies, mineral acids, 
and other poisons which might give a similar test are identified in 
a different manner. 

Carbolic Acid. — Dilute ferric chlorid solution, dilute hydro- 
chloric acid. Add to "x" a few drops of the ferric chlorid solu- 
tion, when a green, blue, or violet color should appear, depending 
on the amount of the poison present. Then add a few drops 
of the hydrochloric acid, when a lemon-yellow color will take the 
place of the blue. Alcohol interferes with this test. Phenol, even 
in very small amounts, has a characteristic odor. 

Chloral Hydrate. — Nessler's reagent. A few drops of Nessler's 
reagent added to a chloral hydrate solution produces a precipitate 
resembling red brick dust, and later this assumes a green color. 
The odor of chloral hydrate is characteristic, reminding one of the 
odor of the American green walnut. Chloral fiends are some- 
times diagnosticated as diabetes patients, as the urochloralic acid 
of the urine reduces alkaline copper solutions. 

Cocain. — Aqueous solution of chromic acid (5-percent) , concen- 
trated hydrochloric acid. To "x" add a few drops of the chromic 



DETECTION OF THE COMMON POISONS. 91 

acid solution. Each drop will produce a precipitate, which will 
immediately disappear if the solution is shaken. Then add to this 
clear solution y 2 dram of the hydrochloric acid. A delicate orange- 
yellow precipitate indicates cocain. 

Copper. — All copper solutions are not blue or green, as is the 
common belief. If, however, clean platinum wire is moistened with 
such solution and held in the colorless flame, the latter will become 
intensely green. 

Ergot. — The chemistry of ergot is still unsatisfactory. Feeding 
ergot to chickens causes gangrene of their combs, but this method is 
of little value when a quick diagnosis is desired. 

Formalin. — In amount sufficient for suicidal purposes, the odor 
is too characteristic to be mistaken, and experimental evidence 
shows that a large amount is necessary to kill. For its detection in 
milk see Milk and Its Home Modifications, page 13-4. 

Hydrocyanic Acid. — The odor of hydrocyanic acid resembles 
that of peach blossoms, and is too characteristic to render necessary 
further means for identification. The poison is very volatile, and 
must be tested at the earliest moment. It should not be inhaled 
in appreciable quantities. 

Lead Acetate (Authors' Method). — Dilute nitric acid, concen- 
trated sulphuric acid. Sugar of lead has a sweetish, astringent 
taste. It shows a tendency to form a milky solution, and contains 
undissolved crystals. Add just enough of the dilute nitric acid 
to clear the cloudy solution, and then add 1 or 2 drops of the con- 
centrated sulphuric acid. The heavy white precipitate is lead 
sulphate. 

In case "x" can not be cleared by a few drops of the dilute 
nitric acid, add 1 drop of the latter to another sample of the 
unknown, filter, and test this filtrate with the sulphuric acid. 

Barium and strontium may respond to the same test, but usually 
need net be considered. In case their presence is possible, barium 
will impart to the colorless flame a yellowish green and strontium 
will show a brilliant red. 

Lead in Drinking Water. — (See Some Simple Water Analyses, 
page 140.) 

Mercury Salts. — Aqueous stannous eWorld solution. Add to 
"x" some of this reagent. A gray and white precipitate indicates 
mercury, being a mixture of quicksilver and calomel. The test 
depends on precipitate formation. Many of these precipitates are 



92 LABORATORY METHODS. 

alcohol soluble, and one should be mindful of the possible presence 
of alcohol in all suspected material. For example, to prove that 
the stomach washings of a drunkard contained corrosive sublimate, 
it might be necessary to heat them slightly to cause the evaporation 
of the alcohol, or else to add an excess of the tin solution. 

Morphin and Opium. — Ferric cJdorid solution. Add to "x" a 
few drops of this reagent. If the color is obscured by a precipi- 
tate, filter it. A deep-blue color indicates pure morphin, while the 
presence of opium is shown by a beautiful dark-red. In case there 
is any doubt, compare with a filtered solution of "x," with the 
reagent, or with controls. 

Phosphorus. — Suicide with match heads has become obsolete. 
At least four are necessary to produce death in an adult, unless 
oils or fats have been taken. The odor and phosphorescence in 
the dark are characteristic. It has been claimed that some rat 
poisons contain phosphorus in addition to arsenic. 

Silver Nitrate. — This turns black when exposed to light. It is 
precipitated from its solution by common salt as insoluble silver 
chlorid. Lead or mercury may answer this test, but either may be 
otherwise identified and differentiated. 

Strychnin. — Concentrated sulphuric acid, crystal of potassium 
bichromate. On a white background — a piece of paper — lay a 
clean slide. On this dissolve some of "x" in 2 drops of concen- 
trated sulphuric acid, and in this mixture crush with a glass rod 
a crystal of potassium dichromate. A beautiful violet or blue 
color indicates strychnin. Very small amounts of the drug are 
bitter to the taste. Some of the sample injected with a hypodermic 
needle into a frog or other animal may bring on the typical tonic 
convulsions. 

Sulphonal, Trional, Etc. — Either drug heated in a dry test tube 
with powdered wood charcoal develops a characteristic odor — 
ethyl mercaptan. If this odor is not familiar to the physician, he 
should run a control test, using sulphonal instead of "x. " 

Less Frequently Applied Tests. — Other substances, the descrip- 
tions of whose detection have been left to larger books, may be 
named as follows : anilin compounds, nicotin, aconite, veratrin, 
codein, oxalic acid, santonin, so-called ptomain poisonings, etc. 

Difficulties and How to Avoid Them. — One of the most com- 
mon causes of poor results in chemical analyses is working with 
the wrong reagent, and this is not always the fault of the worker. 



DETECTION OF THE COMMON POISONS. 93 

Too many registered pharmacists have forgotten their chemistry, 
and examination of certain solutions supposed to be standardized 
by an expert pharmacist showed that no such work had been done. 
One of the authors was unable to obtain the diazo test in four cases 
which he knew to be typhoid, and an examination of his sulphanilic 
solution showed a 50-percent sulphuric acid. Too much must not 
be taken for granted. 

Nomenclature is often confusing. Neither chromic acid nor car- 
bolic acid is properly named. Glycerin is constantly referred to 
by some druggists as a sugar, but is really an alcohol. Quicksilver 
contains no silver, but is metallic mercury. These terms have not, 
however, been avoided in this book, as their use is too widespread. 

A case is known where, in testing for lead by the iodides, a 
worker, usually careful, thoughtlessly employed lead iodid. For 
cautions against explosives and incompatibilities, see Laboratory 
Prophylaxis, page 180. 

It is impossible in a book of this scope to teach the principles of 
chemical analysis, but the following general rules may prove of 
value to the man expecting good results : 

1. Absolute cleanliness. Test tubes can not be kept too clean. 
Sugar of lead is especially prone to adhere to glassware. Do not 
touch with your fingers chemically pure reagents, but pour directly 
from their bottles into solvent or on clean paper, and then cork 
the bottle tightly. Never lay a cork or stopper on a table, but hold 
between fingers until it can be replaced. 

2. Allow time for a reaction to take place before adding more 
reagent or deciding on a negative result. It can not always be 
explained why the same test may at different times employ more 
time. 

3. If precipitates interfere with a color reaction, filter and ex- 
amine the filtrate. 

4. Before performing the Marsh test, study well the properties 
of hydrogen, how it is generated, and how handled. 

5. Unless a worker is certain of his technic, a control test should 
be used in connection with each investigation. Many shortcom- 
ings may thus be detected which would otherwise escape observa- 
tion. 

Value and Limitation of These Investigations. — These have been 
pointed out in the matter preceding the tests. 



CHAPTER VIII. 
EXUDATES IN BRIEF. 



Apparatus. — 

1. Centrifuge. 

2. Eosin. 

3. Exploratory needle. 

4. Formalin. 



5. Glycerin. 

6. Microscope and accessories. 

7. Potassium hydrate. 

8. Sodium citrate. 



Obtaining an Exudate. — An ordinary hypodermic outfit may 
sometimes be used, but it is advisable to employ a long and stout 
exploratory needle. All apparatus should be sterile, especially if 
any cultures are attempted. A small amount of the fluid suffices 
for the tests described in this book, but, as larger amounts are often 
removed for therapeutic purposes, it would not be a waste of time 
to take the specific gravity, or even attempt other examinations 
described in the larger volumes. A bacteriological culture may be 
easily made, but usually is not necessary for safe conclusions. If 
apparatus is sterile, any germ may be looked upon with suspicion, 
but the authors have often been able to gain more from a study of 
the cells than from a search for specific microorganisms. A safer 
plan is to investigate both. An ethyl chlorid spray will render 
these operations painless. 

Thoracic Puncture. — The spot selected for pleuritic effusions is 
marked by the junction of the axillary line with the seventh inter- 
costal space, and is clearly shown in Fig. 25. The arm is raised and 
the hand placed on the opposite shoulder in order to widen the 
interspaces. The relation of the intercostal artery to the lower edge 
of the rib must be kept in mind. Pericardial puncture is rarely 
attempted outside of the hospital. The technic may be found in the 
larger books. 

Abdominal Puncture. — The spot varies, but is usually low. A 
short needle is desirable, so that the bowels may not be punctured. 

Lumbar Puncture. — A short, strong sterilized needle is employed, 



References. — Gruner: Puncture Fluids; all works on clinical diagnosis, including 
Ralili, W'>od, Boston, Simon, etc. 

94 



EXUDATES IN BRIEF. 



95 



but no syringe should be used unless its plunger has been previously 
drawn. The patient should lie on the left side, with the thighs 
flexed and shoulders bent forw T ard. A line joining the iliac crests 
passes between the third and fourth lumbar vertebra. Select a 




Fig. 25. — Puncture of pleura. 



point midway between the spinous processes of these two bones, 
and place the point of the needle one-third of an inch to the left 
of this point and insert. Fig. 26 illustrates the exact position. 
Allow the liquid to flow as it will, and do not aspirate. If fluid 
does not come, have the patient strain, and it may be necessary to 
again insert the needle. The operator should be certain that the 
needle is not clogged before attempting this procedure. In these 
punctures, as well as all others including thoracic, abdominal, 
joints, etc., several precautions are emphasized : 
1. All apparatus must be sterile. 



96 



LABORATORY METHODS. 



2. Avoid important structures, as nerves, vessels, intestines, blad- 
der, heart, etc. 

3. Do not aspirate in lumbar punctures. If necessary to carry 
out this procedure elsewhere, do so very slowly and then permit ex- 
cess to drain. 



^^W^M^^^^^ 




m\\\\v-^^r-- 






Pig. 26. — Diagram showing site of lumbar puncture. x,x, tips of spinous processes; x' 
point midway between ; A, lower tip of spinal cord; B, ilium. 



4. If patient shows any signs of collapse, stop the operation at 
once. Very often these punctures may be followed by a prompt 
relief of symptoms. 

Preparing an Exudate. — As cells and bacteria may be difficult to 
find, it is usually advantageous to centrifugalize as with urine. 
Many of these exudates show a tendency, however, to clot very 
quickly, and to avoid this the tube may be one-quarter filled with 
2-percent sodium citrate solution. The following process may then 
be followed : 

i. Centrifugalize sufficiently. 

2. Remove with a pipette the supernatant citrate solution. 

3. Add an equal amount of dilute formalin, 1-percent aqueous 
solution. 



EXUDATES IN BRIEF. 97 

4. Mix thoroughly by shaking. 

5. Permit to stand five minutes. 

6. Centrifugalize sufficiently. 

7. Remove sediment from beneath liquid by the method de- 
scribed for urinary sediments. (See The Urine in Disease, 
page 116.) 

8. Make spreads, dry, and fix by heat ; stain Avith methylene blue. 
Most Important Findings. — Various chemical tests can hardly 

be considered here. The cytoanalysis is doubtless the most im- 
portant, and may be briefly given as follows : 

1. Tuberculosis is characterized by lymphocytes and few red 
cells. 

2. Tuberculosis with secondary purulent infection is character- 
ized by a mixture of lymphocytes, polymorphonuclear leukocytes, 
and red cells. 

3. An infection never tuberculous, but caused by the so-called 
purulent germs — as gonococcus, meningococcus, pneumococcus, 
streptococcus, colon or typhoid bacilli, etc. — is characterized by 
absence of red cells and lymphocytes, but large numbers of poly- 
morphonuclear leukocytes are present. 

4. Ascites, etc., contains a few mesothelial cells — large flat cells, 
with single nuclei. 

Searches may be made for the various microorganisms. In case 
fluid from the lumbar puncture shows lymphocytosis, especially 
without red blood cells, the process may be syphilitic instead of 
tuberculous. Age, history, and other factors must be taken into 
consideration. If, however, it contains only polymorphonuclear 
leukocytes, and these in considerable numbers, it may be a menin- 
gococcus infection; in fact, this organism may be detected within 
the pus cells. Meningitis of an acute type has been caused by 
other germs, notably the pneumococcus, gonococcus, and typhoid 
bacillus. A few white cells may be found in a normal serous fluid. 

Less Frequently Applied Procedures. — The practitioner does not 
usually attempt the diagnosis of general paresis by lumbar 
punctures. If he desires to "tap" the pericardial cavity, complete 
descriptions of this procedure may be found in the larger books. 
Fluids from ovarian cysts, spermatocele, etc., offer little to the 
diagnostician. 

Azobspermatism. — Family sterility is due in about two-fifths of 
all cases to lack of spermatozoa in the male. The condition of azo- 



98 LABORATORY METHODS. 

ospermatism can not be diagnosticated by a single examination, or 
by less than a dozen such investigations, to be made monthly. 
Spermatozoa may be absent from the semen during certain dis- 
eases. The frequent exposure to x-rays may cause a destruction of 
all spermatozoa and result in a permanent sterility. Spermatozoa 
may be present, but may be dead at the time of emission, falling 
a prey within the male to the treponema or other injurious influ- 
ences. Such germ cells are not motile, but, in order to prove that 
loss of motion is not due to chilling, examination must be made at 
once, using warmed slides and cover glasses. 

Spermatorrhea. — The male germ cells may be constantly present 
in the urine of many men, and is caused by certain sexual excesses, 
masturbation, and severe diseases. The urine of the female may 
contain spermatozoa, which exist as a vaginal contamination after 
intercourse. 

Proof of Rape. — So far as legal processes are concerned, such 
questions are left to experts. The parents of the girl may, how- 
ever, desire a private opinion, and they may not wish to subject 
the girl to an examination, but may bring some vaginal secretion 
or scrapings, which may be examined at once for spermatozoa; or, 
more commonly, a dry and stained cloth may be submitted. In 
case the semen was pure and is contaminated by little of the vaginal 
secretion, the stain will be almost colorless, and will give the cloth 
a "feel" not unlike that of starched, but unironed, linen. The 
characteristic odor may be present. 

Place a small piece of the stained cloth in a few drops of a 1-per- 
cent aqueous solution of caustic potash, and allow to soak for at 
least two hours. Pour off liquid and add several drops of gly- 
cerin. Tease thoroughly, but gently, as the tails are very easily 
broken. Add 1 drop of eosin and mix thoroughly. Examine some 
of the liquid under oil immersion lens. The spermatozoa are 
stained red, but cotton and linen fibers are unstained. Other cells, 
especially erythrocytes, take this stain, and should be identified 
if they are present. They may have been destroyed in the soaking 
process. A vigorous search should be made for the tails of the 
spermatozoa. 

Vaginal Blenorrhea. — Noninfectious leucorrhea may be caused 
by sexual excesses, dysmenorrhea, and many other conditions not 
uncommon to feminine physiology, as well as by tumors, malposi- 
tion of the uterus, etc. So far as macroscopical findings are con- 



EXUDATES IN BRIEF. 99 

cerned, this discharge may resemble true pus; microscopically it 
may contain polymorphonuclear leukocytes, and then a vaginitis 
may be diagnosticated. 

Purulent Leucorrhea. — Under this head are not usually included 
the nonspecific forms of vaginitis, but only those in which the 
gonococcus may be demonstrated. (See Searching for Germs, 
page 48.) 

Dental Caries. — In certain chronic diseases and in fevers the 
saliva may become acid, and at other times an alkaline reaction may 
occur. A persistent acidity is usually due to lactic acid, which is 
formed by certain bacteria, which often results in cavity formation. 
Phenolphthalein solution rather than litmus should be used as an 
indicator. 

Mercurial Ptyalism. — Salivation may be due to many causes. 
Mercury may be detected in the saliva according to tests described 
on page 91. 

Noguchi's Butyric Acid Test for Syphilis. — Noguchi found that the 
active lipotropic bodies causing the Wassermann reaction were con- 
tained in or at least precipitated with globulin ; and that the globulin fraction 
is increased in syphilis. To detect this increase, he has devised the butyric 
acid test. This may be applied to the cerebro-spinal fluid as follows: — In a 
very small test tube, 3 or 4 drops of this fluid are mixed with ten drops of 
a ten-percent butyric acid .solution in physiologic salt solution. This mix- 
ture is heated over a flame and is boiled for a brief period. Two drops of 
a normal solution of sodium hydrate are then added quickly to the heated 
mixture, and the whole boiled once more for a few seconds. The cerebro- 
spinal fluid employed must be free from blood. 

Within two hours, a white and granular precipitate should appear which 
will settle to the bottom of the tube. The greater the amount of protein, 
the more quickly the reaction will occur. Any precipitate which forms after 
two hours may be ignored. 

Interpretation of the Butyric Acid Test. — This reaction, though 
valuable in the diagnosis of the syphilitic and parasyphilitic affec- 
tions, may occur in any of the acute inflammatory conditions of the meninges 
and in tuberculous meningitis. However all of these may be readily differ- 
entiated at least from the parasyphilitic affections where acute symptoms 
and fever are usually absent. The butyric acid test has been found to be 
present in poliomyelitis. Normal cerebro-spinal fluid may give a slight tur- 
bidity but never a granular precipitate within two hours. The test is 
scarcely as delicate as the Wassermann in its positive phase; but the nega- 
tive butyric acid test in the diagnosis of syphilitic and parasyphilitic condi- 
tions, is much more reliable than the negative Wassermann or Perutz (see 
page 199) : the former excludes syphilis, but the latter do not. In case the 
Perutz test is negative, the butyric acid test should be carried out. A nega- 
tive butyric acid test almost excludes the possibility of true tabes or general 
paralysis. 

The reaction is also of value in differentiating between the various forms 
of acute meningitis and the other infectious diseases. Thus in typhoid, 
typhus, malaria and the exanthemata, the reaction is negative. 



CHAPTER IX. 

DIAZO VERSUS WIDAL. 

This chapter is designed to set aright the puzzled practitioner, 
and it is probable that many of the recommendations would hardly 
find favor with hospitals or college clinics. Were it not for the 
purpose of comparison, these two investigations would be included, 
respectively, in The Urine in Disease and Searching for Germs. 

EHRLICH'S DIAZO REACTION. 

Advantages in the Bedside Diagnosis of Typhoid. — These ad- 
vantages are as follows : 

1. It occurs early — that is, when a final diagnosis is important. 
It is expected during the first week, and it is often seen from the 
third to the fourth day after the onset. It is commonly observed 
before the rash appears (Osier). The authors have noted its pres- 
ence in cases not yet bedfast — that is, during that period of lassi- 
tude, headache, and chilly feeling which so of ten precede the onset 
proper — and patients with such symptoms have been sent from the 
consultation room to the sickbed with a fairly safe diagnosis. 

2. The diazo test may be easily and rapidly completed at the 
bedside, requiring three solutions, a test tube, and three minutes ' 
time. 

3. The reaction is present in almost every case of typhoid. 

4. The reaction is, in a practical sense, specific. It occurs in 
other diseases, but, with the exception of a few cases of miliary 
tuberculosis, these should rarely be confused with typhoid. In 
these cases of acute miliary tuberculosis the reaction is rarely, if 
ever, present during the first week. 

5. The diazo reaction may differentiate between a relapse and a 
complication when all other signs fail, and it often reappears be- 
fore or is coincident with recurrence of the fever, but is not ob- 



References. — Sahli, Emerson, Simon, Wood, Boston, and other works on clinical 
diagnosis. 

100 



DIAZO VERSUS WIDAL. 101 

served in connection with appendicitis, perforation of the gut, or 
in iliac thrombosis. 

Disadvantages of the Diazo. — Ehrlich's test is not without its 
shortcomings, as here noted : 

1. It is not absolutely pathognomonic. 

2. Unfortunately the diazo reaction is not always present. A 
negative test does not imply that the disease is not enteric fever, 
but the percentage of cases in which the diazo is really absent must 
be small indeed. 

Technic of the Diazo Reaction. — The following apparatus and 
reagents are necessary. 

Solution 1 — sulphanilic acid ; saturated solution in 5-percent 
hydrochloric acid. 

Solution 2. — sodium nitrite; ^-percent aqueous solution. 

Solution 3 — aqueous ammonia. 

Test tube. 

Medicine dropper. 

The three solutions may be obtained from reliable manufacturers. 
Put 51 drops of urine in a test tube and add 50 drops of solution 
1 and 1 drop of solution 2. Shake thoroughly. Then add quickly 
about 5 drops of ammonia, allowing it to run down the side of the 
test tube in such a manner as to form an upper layer. If the test 
is positive, a deep-pink or rose-red colored ring will appear at the 
junction of the liquids. Shake the mixture. The entire bulk and 
also the foam should become red, the coloration of the foam being 
most characteristic of the reaction. If the urine is allowed to 
stand until the next visit, a dark bile-green sediment may be 
present, and, while of value, its absence does not indicate a nega- 
tive test. The authors observed one case in which this precipitate 
was formed immediately on the addition of the ammonia and be- 
came very dense after one-half hour. A pink color which formed 
at first was not detected fifteen minutes later. The patient was 
delirious at the time, but eventually recovered. No satisfactory 
explanation of this phenomenon was ever proposed. 

Pseudo-Diazo Reactions. — In health, salmon- or orange-colored 
rings may be observed, but not the characteristic pink or rose-red 
colors. The foam is not colored, and the green precipitate fails to 
form. Where large amounts of urine are passed, the causative 
factor may be so diluted that a positive reaction is obtained with 
difficulty, or not at all. 



102 LABORATORY METHODS. 

Salol, betanaphthol, and opium, when administered, may cause a 
pseudo-reaction. The foam is not, however, pink or red, but is 
colorless or yellow, and the green precipitate never forms. Bis- 
muth subgallate and the tannic acid derivatives prevent the posi- 
tive diazo reaction from appearing, and it is due largely to failure 
to take this into consideration that the total percentage of positive 
tests is held lower than is really just. If jaundice is present, a 
dark, cloudy discoloration may obscure the true reaction. 

In certain cases of pneumonia, which may or may not simulate 
typhoid, a yellow color may appear on the addition of solution 2. 
Ammonia changes this to a lemon tint. Such a reaction, "egg 
yellow, ' ' is said to have been observed in cases of typhoid fever. 

In certain affections, as erysipelas, rheumatism, and other dis- 
eases not easily confounded with true typhoid, the true diazo has 
been observed. 

Modifications or Substitutions for Diazo Reaction. — These are 
listed, but not described in detail. 

1. Greene's Modification. The test is said to be more delicate 
when double the amount of solution 1 is used. With this modifi- 
cation the same amounts of the other reagents are used. 

2. Friedenwald 's Suggestion. Paramidoacetophenon is sub- 
stituted for the sulphanilic acid. 

3. Ehrlich's Modified Diazo. Where dimethylaminobenzalcle- 
hyde and other reagents are employed. (See page 114.) 

4. Russo's Reaction. 1 Where methylene blue is used. 
Sources of Error in Technic. — These are few, but important. 

The instructions, if followed minutely, may be depended upon. 
One of the most frequent causes of difficulty may be traced to 
adding the ammonia layer too slowly. When this occurs, the re- 
action may not take place. Other sources of error may be recog- 
nized by a careful study of the pseudo-reactions. 

Value and Limitations of the Diazo Test. — From what has been 
said, the value of this reaction may be summed up about as follows : 
With certain limitations, it is especially useful as a bedside diagnos- 



1 This is a simple and seemingly valuable method of differentiating between typhoid 
fever and acute mjliary tuberculosis. The urine of the former will be turned a beau- 
tiful emerald color when 4 drops of a 1:1,000 aqueous solution of methylene blue are 
added to 4 or 5 cc. of the sample. In the latter disease only a bluish or greenish 
tinge is obtained. It is advisable to use a normal urine in a control test if there is 
any question in regard to the color changes. As the positive reaction occurs in other 
pathological conditions, it is valuable only in the differentiation named, and should 
always be used in connection with the diazo. 



DIAZO VERSUS WIDAL. 103 

tic procedure, and these circumscriptions have been emphasized 
under disadvantages, errors, pseudo-reactions, etc. 

WIDAL REACTION. 

Widal Reaction Defined. — "While typhoid agglutination tests in 
general are considered, the technic described will deal only with 
the suspensions of dead cultures and with macroscopic observations.^ 
These seem to be not only sufficiently reliable procedures, but are 
specially adapted to bedside work, and the original test could hardly 
be made except with the equipment of the large laboratory. 

Advantages of the Widal Test. — These advantages are as fol- 
lows : 

1. It is pathognomonic of enteric fever, thus differentiating it 
from acute miliary tuberculosis. 

2. May occur in those few cases where the diazo test is absent, 
though not usually so early. 

3. Its appearance, even if too late for diagnosis so far as treat- 
ment is concerned, serves to complete hospital records, death cer- 
tificates, vital statistics, etc. The reader must not infer that the 
Widal is invariably a late reaction. 

Disadvantages of the Widal Test. — These disadvantages are as 
follows : 

1. It is of little or no prognostic value because it does not vary 
or reappear with a relapse, and can not therefore differentiate it 
from serious complications. 

2. Takes too much time to perform. Even with the simple tech- 
nic described, a diagnosis must be delayed for at least a few hours 
— a long time when we consider that we are dealing with an acute 
disease. 

3. Although sometimes present on the fourth day, it usually 
appears late. Some observers (Stitt and the authors) do not 
recommend it as a routine procedure until the second week. 

4. It is not always present in true typhoid. Although it usually 
appears at some stage of enteric fever, its invariable presence in 
this disease has yet to be proven, and a negative reaction does not 
imply no typhoid. So far as percentage is concerned, one thing 
is certain — during the early manifestations of the disease, when 
diagnosis from a therapeutic standpoint is most important, the 



104 LABORATORY METHODS. 

diazo reaction occurs in much higher proportion of all cases than 
does the agglutination test. 

Technic of the Widal. — The general practitioner should appre- 
ciate the efforts of certain manufacturers in providing the simpli- 
fied Widal test, but the authors do not recommend the expensive 
outfit. The manufacturers are usually willing to supply the dead 
cultures of the typhoid bacilli, properly preserved and suspended, 
without the fancy pipettes, etc. The remainder of the apparatus 
consists of two homeopathic vials. 

The technic is not complex. Add about 1 dram of the suspen- 
sion to each vial. One of these is corked and serves as a control. 
Into the other add 2 or 3 drops of the patient's blood directly 
from a finger prick. Shake and cork. Set aside both vials and 
examine in several hours. These bottles can not be carried in the 
pocket without interfering with the test, so that, unless the physi- 
cian's office is near, a return call is necessary. Caution the family 
not to touch the vials. A positive test is observed when the sus- 
pension becomes granular or milky and the germs sink to the 
bottom, leaving an upper clear fluid. The control should remain 
cloudy. 

Bass and Watkins' Rapid Method for Widal. — The blood is 
diluted by dissolving it in approximately four times its volume 
of water. Then one or two drops of this diluted blood are mixed 
on a microscopic slide or other piece of glass with an equal quan- 
tity of the suspension of dead typhoid bacilli. The slide is tilted 
from side to side or end to end in order to keep the mixture 
flowing back and forth. If the reaction is positive, a grayish- 
mealy sediment appears within one minute, usually much more 
quickly. This consists of agglutinated bacilli and is easily seen 
with the unaided eye. It appears in the fluid around the edges 
first, and tends to collect there. If the agglutination is contin- 
ued, the clumps increase in size for two or three minutes. "With 
blood giving a weak reaction, the appearance of the sediment is 
not so rapid as with stronger reacting blood. It is useless, how- 
ever, to continue the test longer than two minutes, for, if the re- 
action has not occurred within that time, it will not ensue at 
all. When the reaction is negative, no agglutination occurs ; and 
the mixture remains as clear and unchanged as when placed upon 
the slide. The suspension is best made up containing ten thou- 



DIAZO VERSUS WIDAL. 



105 



sand million dead typhoid bacilli per cc. in 1.7 percent sodium 
chloride solution to which one percent of formalin has been 
added. Keep well stoppered in an amber bottle and in a dark 
place. 

Table of Comparisons. — It will be observed that this chapter 
has been written for the needs of the general practitioner, and the 
comparative table given below has been prepared with the same 
object in view. Typhoid is an acute and dangerous disease. Cor- 
rect therapeutic measures must be instituted promptly, and delays, 
unless made necessary by submitting specimens to distant labora- 
tories, must be avoided. The technic and significance of these re- 
actions are so simple that the attending physician has only him- 
self to blame if his patient dies "waiting for a diagnosis." 



DIAZO. 

1. Appears early in typhoid fever. 

2. Reappears with relapses, but not 

with complications. 

3. Technic rapidly completed. 

4. Does not differentiate acute mil- 

iary tuberculosis; when delayed 
until the third week, suggests 
tuberculosis of a general type. 

5. Not always present. 

6. Occurs in other diseases not easily 

confused with typhoid. 



WIDAL. 

1. Appears in typhoid fever. 

2. Relapses do not influence the test. 

3. Technic in simple form takes sev- 

eral hours. 

4. Not present in acute miliary tu- 

berculosis. 



5. Not always present. 

6. Occurs in no other disease. 



The authors have no intention to detract from the Widal test, 
but simply desire to classify it properly and then to emphasize 
those advantages of the diazo reaction usually overlooked or neg- 
lected. The physician may make both tests, but should not omit 
the diazo. 

Less Frequently Applied Procedures. — Both the microscopic 
and macroscopic tests with the living bacilli are best conducted by 
the larger laboratories, but, in spite of emphatic statements to the 
contrary, these seem to possess no special advantages over the test 
here described. 



106 



LABORATORY METHODS. 




CHAPTER X. 



THE UKINE IN DISEASE. 

Apparatus. — Does not include that used in the urobilinogen 
test. 



1. Nitric acid, concentrated. 

2. Centrifuge or sedimentation 

glasses. 

3. Ferric clilorid solution, strong. 
•4. Graduate. 

5. Haines' solution. 

6. Lead acetate solution, strong. 

7. Medicine dropper. 

8. Microscope and accessories. 



9. Phenolphthalein, 1 -percent alco- 
holic solution. 

10. Pipette, long. 

11. Silver nitrate solution, 15-percent. 

12. Saccharimeter (Einhorn). 

13. Squibb's urea apparatus. 

14. Urinometer. 

15. Window glass, 2x2 inches. 



Scope of This Chapter. — It has been the aim of the authors to 
select the best tests, and to describe them simply, but thoroughly, 
with a view to assist the physician who must do his work hurriedly 
and who must often search through dozens of methods to find the 
one which he may properly apply without laboratory luxuries and 
one npon which he may safely depend. In many cases a diagnosis 
without urinalysis is valueless, but, on the other hand, it may be 
stated that " conclusions drawn from the water alone are as brittle 
as the urinal containing it." 

Amount, Significance, Etc. — The normal amount varies from 30 
to 45 ounces every twenty-four hours, and the normal and patho- 
logical variations bear the following indications : 

Increase of Urine. Due to cold, diabetes, amyloid kidney, inter- 
stitial nephritis, some brain tumors, and following fever. 

Decrease of Urine {not Including Anuria). Due to hot weather 
and perspiration, parenchymatous nephritis, diarrhea, hysteria, 
and during fevers. 

Specific Gravity, Significance, and Estimation. — This varies 
normally between 1.012 and 1.025, being high or low respectively 



References. — Memminger : Diagnosis by the Urine; Tyson: Examination of the Urine; 
Saxe: Examination of the Urine; Rieder : Urinary Sediments; all works on clinical 
diagnosis, especially Sahli, Boston, Simon, Wood, etc. 

107 



108 LABORATORY METHODS. 

as there is little or much urine passed daily. In parenchymatous 
nephritis it is high, and there is little urine; in diabetes there is 
polyuria, but a high specific gravity ; in insipid diabetes and inter- 
stitial nephritis there is a persistent polyuria, with a low specific 
gravity. 

Estimation of the specific gravity by means of the urinometer, 
when carefully done, is an entirely different procedure from that 
usually conducted even in a perfectly arranged laboratory. A 
correct reading is impossible where the following precautions are 
overlooked : 

1. All foam should be taken up from the surface of the urine 
by means of a piece of filter paper or blotter before the bulb is 
immersed. 

2. The urinometer should float freely toward the center of the 
sample, and should not be in contact with the inner surface of the 
cylinder. 

For practical purposes, temperature corrections are not neces- 
sary, providing the sample is neither very cold nor has been 
heated. 

Color, Significance. — The color of normal urine varies from that 
of copper to brass, and the color changes bear the following indi- 
cations : 

Colorless urine — polyuria ; red urine — fresh blood, urates ; brown 
urine — blood, urates ; smoky urine — blood ; gray urine — pus, al- 
bumin ; blue urine — indican, methylene blue ; black urine, on stand- 
ing — alkapton formed normally or after the administration of aro- 
matic series of carbon compounds, the color usually first appearing 
at the top of the sample and proceeding downward. 

Odor, Significance. — There is a characteristic normal odor due 
to certain volatile organic substances. Urotropin administration 
imparts to the urine an odor of formaldehyd. The sweet odor of 
diabetes needs no comment. A urine, standing, gives off odors 
characteristic of alkaline fermentation (page 121, 122). 

Transparency, Significance. — A normal urine is usually clear, 
but may contain urates when voided, especially after exposure to 
cold, and may contain amorphous phosphates after a large meal. 
A slight cloudiness may be due to those small quantities of mucus 
and cells normally present. 

Marked cloudiness is usually pathological to the fresh urine. If 
due merely to salts, a little heat and a drop of nitric acid will clear 



THE URINE IN DISEASE. 109 

it. Pus, albumin, and blood do not disappear during the process, 
and their identification must be accomplished by other tests. 

Reaction, Significance, and Determination. — A slight acidity is 
normal. A slight alkalinity can not, in many cases, be termed ab- 
normal. Normal acidity is not dependent en, nor perhaps influ- 
enced by, uric acid, but is caused by certain phosphates. 

Marked alkalinity of freshly voided urine may suggest cystitis, 
but alkalinity of a decomposing urine has no clinical significance. 
A more complete treatment of this subject is found under the com- 
parison of acidemia and acidosis (page 123). 

The ordinary litmus test is worse than a waste of time in prac- 
tical urinalysis. Acidity versus alkalinity may be first solved by 
the more delicate indicators, and these serve not only to draw finer 
distinctions, but actually give some idea as to the degree of the 
reaction. The authors use a dilute solution of phenolphthalein, 
such as is employed in the gastric analyses. In acid or neutral 
media this solution is colorless, but, if one drop is added to an 
alkaline solution, a prompt color reaction takes place, varying from 
a light-pink to a deep-crimson, depending on the degree of alka- 
linity. A simple method of estimating urinary acidity is as fol- 
lows: 

Add to the urine a drop of the indicator. A colorless solution 
signifies that the urine is either neutral or acid. Now add a drop of 
the sodium hydrate solution used when determining the percentage 
of total acid in the gastric juice (Chemistry and Biology of the 
Gastric Juice, page 76). If the reaction was neutral, a pink or red 
color will appear. The acidity may be determined approximately 
in degree by the number of drops of the alkaline solution necessary 
to impart to the sample a permanent red color, and this permanency 
can be tested by stirring thoroughly. In case exact readings are 
desired, titration may be conducted as described in Chemistry and 
Biology of the Gastric Juice (page 76). 

Significance of Albumin. — The sources of urinary albumin are 
many and its etiology varied, but albumin is not expected to be 
found in a freshly voided normal urine. The following list will 
serve to give an idea of the principal sources of protein bodies : 

Serum Proteins (which may or may not cloud the voided urine). 
1, fevers ; 2, diseases of the blood ; 3, poisonous drugs ; 4, accom- 
panying heart disease ; 5, certain functional and dietetic types ; 
6, organic kidney lesions. 



110 LABORATORY METHODS. 

Urinary Suspensions of Cellular Elements. 1, pus or epithelial 
cells in considerable numbers ; 2, blood cells ; 3, bacteria. 

Choice of Tests. — The heat and nitric acid test — not the "nitric 
acid and heat" test — when properly conducted, often suffices, but 
can not be recommended as the main procedure, nor as the choice of 
methods when only one test is used, as it will not demonstrate 
traces of albumin. The method is, however, valuable, and should 
be included mainly to rule out urates, which are dissolved by the 
heat, and earthy phosphates, which are precipitated by a rise in 
temperature, but disappear on adding the acid. If an efferves- 
cence occurs on adding the acid, it is due to the formation of ni- 
trogen gas or carbonic acid gas, depending upon the reaction of the 
urine. 

It is, however, the trace of albumin that concerns us in diagnos- 
tics, and this small amount, as in diabetes and interstitial nephritis, 
may mean much more to the attending physician or life insurance 
examiner than any other single symptom. The first described 
method is merely a modification of Boston 's pipette method, a medi- 
cine dropper, such as is used by every physician, being substituted 
for the regular pipette. 

Boston's Pipette Method. — Compress the bulb of a clean medi- 
cine dropper (Fig. 28), immerse the point in a sample of the urine 
and slightly relax the pressure on the bulb, when a small quantity 
of the urine will enter the tube. Carefully wipe off the outer sur- 
face of the tube, immerse the point in a small quantity of concen- 
trated nitric acid, and again relax the pressure sufficiently to draw 
up a column of the liquid. If albumin is present, a distinct white 
ring forms at the junction of the liquids. This modification offers 
no advantage over Boston's method, except that of a bedside di- 
agnosis, where a medicine dropper and a bottle of nitric acid may 
be conveniently used. It is more simple, and at the same time more 
likely to show the trace of albumin, than the heat and nitric acid 
method, although both tests should, if possible, be made. 

Heat and Nitric Acid Test. — Fill a clean test tube half full of 
urine and boil, when a cloudiness may result, which may be due 
either to phosphates or to albumin. The addition of a few drops 
of nitric acid will clear the former, but not the latter. In rare in- 
stances, calcium phosphate may be dissolved with difficulty. 

Filtration of the Urine.— Filtration of a cloudy urine is ad- 
visable when searching for traces of albumin, and will remove' all 



THE URINE IN DISEASE. Ill 

cells and most bacteria. In case, however, the physician is com- 
pelled to make the examination at the bedside, he may often omit 
it, comparing the tested sample with one which has not been exam- 
ined, and in this way he will be frequently able to note marked 




Fig. 28. — Modification of Boston's pipette test for albumin. 

differences in the amount of albumin present. In other words, 
while it is advisable to use only a clear urine when testing for 
urinary albumin, the absence of such apparatus should not interfere 
with making the test, and it may be possible to obtain in the house- 
hold an ordinary tin funnel and a piece of paper suitable for the 



112 LABORATORY METHODS. 

procedure. Contrary to the general idea, the laity, as a rule, do 
not discredit these examinations. 

Estimations of Albumin. — It is indeed strange that the trace 
of albumin is often of greater prognostic import than the large 
flaky or putty-like precipitate, and for this matter the reader is 
referred to the larger books for the various albuminometers. 

Sources of Error. — It does not necessarily follow that an al- 
bumin reaction indicates a renal lesion. If the urine has not been 
filtered, bacteria and the various cells may cause the positive test 
to occur. Pseudo-tests may be observed, and should be ruled out 
by the following methods when their presence is suspected : 

1. Urates. These quickly dissolve when heated. 

2. Santal Oil, Copaiba Oil, Etc. A few drops of alcohol added to 
the urine quickly dissolves these or prevents their formation. 

Significance of Glucose. — Glycosuria may be transient or diabe- 
togenous, and before the latter diagnosis can be made the occur- 
rence of this condition must be shown to be persistent, or at least 
to persistently recur even when the carbohydrate food is somewhat 
diminished. The presence of diacetic acid or of the other acetone 
bodies signifies sugar diabetes. In case of doubt, a record of 
twenty-four-hour amounts and specific gravity tests may clear up 
the diagnosis. Glycosuria is merely a sequence of hyperglycemia. 

Detection of Glucose. — The authors recommend Haines' test, not 
merely because it is simple, but because it gives such satisfactory 
results. A small bottle of the reagent may be obtained at any 
pharmacy, and does not necessitate a fresh preparation for each 
test. In time, however, a reddish precipitate will settle to the 
bottom of the bottle, when a fresh supply should be obtained. On 
boiling this reagent, before the urine is added, precipitation should 
not occur. If albumin is present in the sample to be tested, it 
should be removed previous to the examination. The procedure is 
as follows : 

1. Boil in a clean test tube for two minutes 1 dram or more of 
Haines' reagent. 

2. Hold in the light to make certain that no change has occurred. 

3. Quickly add from 2 to 7 drops, not more, of the urine. 

4. Keep the reagent boiling before the urine is added, but do 
not heat after this has been done. 

5. A heavy yellow, brown, or red precipitate indicates the pres- 
ence of glucose. A slight flalrv- white or dirtv collection of crys- 



THE URINE IN DISEASE. 113 

tals indicate merely a partial reduction of the reagent, and do not 
indicate that glucose is present. The typical precipitate is heavy, 
both in appearance and in reality, settling to the bottom of the test 
tube like sand, and, when once seen, is never confused with the 
pseudo-reactions. 

Estimation of Glucose. — This may be necessary in certain 
dietetic tests, but certainly never as a prognostic procedure, the 
increase of the acetone bodies serving the latter purpose. 

The Einhorn saccharimeter offers a method that is simple and 
fairly accurate. Its chief objection is that a wait of twenty-four 
hours or longer is required for its completion, a criticism that 
seems inconsequential when the chronic nature of diabetes is con- 
sidered. 

Sources of Error. — Haines' test is satisfactory when directions 
are properly followed. Glucosides and certain volatile oils, when 
taken internally, are often excreted by the kidneys. When boiled, 
they break up into simple substances, including either free glucose 
or similar bodies, which would, of course, reduce Haines' solution; 
but these will not, however, cause any trouble if the contents of the 
tube are not heated after the urine is added. As noted above, 
albumin will interfere with the test, and should, if possible, be 
precipitated and filtered before testing for glucose. The physi- 
cian may advantageously at some idle moment attempt this test 
with a sample of urine to which a little glucose has been added. 
A qualitative fermentation test, with a little yeast in an ordinary 
bottle, leaves no doubt as to the presence of glucose. 

Significance and Detection of Bile. — Biliary products, such as 
are finally excreted by the kidneys, appear in the urine as uro- 
bilin, and are the cause of its normal yellow or orange tint. In 
jaundice, however, bilirubin and its oxygenated derivatives may be 
present in the urine. The best test for these may be made by 
merely shaking well the sample and observing carefully the color 
of the foam. In normal urine this is white, but, if abnormal biliary 
pigments are present, a yellow, green, or brown coloration will be 
noted. For practical purposes no other test need be attempted. 

Significance of Blood. — Fresh or changed blood may come from 
any portion of the urinary apparatus. It is claimed that renal 
and vesicle hematuria may be distinguished by several mechanical^ 
methods. One of these requires the urine to stand twenty-four 
hours in a conical glass. In vesicle hematuria all blood will have 1 * 



114 LABORATORY METHODS. 

settled to the bottom of the glass, but in the renal forms the entire 
liquid will remain smoky. Fresh blood indicates, most probably, 
a vesicle source. Casts containing blood cells point to renal origin. 

A red urine may be due to fresh blood or to urates. Heating 
the sample causes rapid solution of the latter, but has no effect on 
the former. 

The Urobilinogen Test for Hepatic Function. — Neubauer and 
others have shown that the Ehrlich's modified diazo reaction (see 
page 102) is not after all a test for typhoid fever, but often oc- 
curs here and elsewhere inasmuch as the liver when involved 
sometimes permits urobilinogen to escape unchanged into the 
urine ; and it is this substance which gives the test. It would 
thus seem that we have found a very valuable as well as ex- 
ceedingly simple method of determining whether or not the 
" liver function " is normal; and this may be of considerable 
diagnostic profit. Thus, for example in the cirrhoses, where most 
of the cells of the liver are simultaneously involved, this organ 
may be unable to "work up" the urobilinogen. In other words, 
so many cells are injured that the remainder are unable (by 
virtue of their inherent possibilities to undergo compensatory 
hypertrophy or other change) to complete the work expected of 
that organ. The test appears to be a very sensitive one, but 
never occurs in perfect health. It appears to be of great value 
in the diagnosis of those liver ailments where the greater part of 
the organ is affected; viz., passive congestion, cirrhosis, biliary 
obstruction, severe fevers with involvement of this organ, hepatic 
syphilis and angiocholitis. Minor hepatic troubles affecting 
most of the cells may give rise to urobilinogenuria whereas se- 
vere diseases of that organ affecting only a small portion of it 
may cause no such condition because enough cells are left un- 
injured to assume the whole work laid out for that cell com- 
munity. 

Do not heat the urine or the reagent, as the results will be 
confusing. Two drops of the testing solution are added to one 
dram of the urine. The mixture is shaken and set aside at room 
temperature. If urobilinogen is present, a beautiful cherry-red 
color will develop after a time varying from a minute to a cou- 
ple of hours. A yellowish or pinkish tint cannot be regarded as 
positive. The testing reagent may be made up and kept indefi- 
nitely. It is prepared by dissolving one gram of para dim ethyla- 



THE URINE IN DISEASE. 115 

minobenzaldehyde in 10 cc. of pure hydrochloric acid and then 
adding five drops of alcohol and enough distilled water to make 
up 50 cc. total quantity. 

Diazo Reaction. — (See Diazo Versus Widal, page 101.) 

Significance of Urea Content. — Normally, about 500 grains of 
urea are passed daily, but this output varies under physiological 
conditions. It is only when the urea runs very low that a patho- 
logical significance may be applied. In certain forms of Bright 's 
disease, and in pregnancy where eclampsia is feared, its estimation 
may be att&npted mainly as a prognostic procedure. 

Estimation of Urea. — Any method requiring the making up of 
fresh quantitative solutions for each estimation finds small favor 
with the practitioner. The Squibb apparatus requires but little 
of this work, avoiding at the same time the use of the irritating 
bromin. This apparatus is sold by instrument dealers, and is ac- 
companied by full directions for using. 

Significance of Uric Acid. — In small amount, uric acid is nor- 
mal, but in increased quantities — due to unknown causes — it may 
lead to the formation of urinary calculi, to gout, and to other forms 
of arthritis. 

Detection of Uric Acid. — In a cold urine, uric acid and its 
homologues appear as a brick-red precipitate. A little heat will 
cause their solution, thus differentiating them from blood. 1 A 
simple test for one of the compounds accompanying the uric acid 
proper may be made by adding 1 or 2 drops of a sugar of lead 
solution to the sample, when a flesh-colored precipitate will be 
formed. A microscopical examination may reveal in the cold urine 
characteristic uric acid crystals. 

Estimation of Uric Acid. — That true lithemia occurs, and that 
an increase of uric acid in the secretion of the kidney proves its 
existence, are not unquestioned medical truths. This statement 
does not refer to true gout, but to the many ailments usually 
attributed to this normal urinary salt, and for these reasons the 
estimation of uric acid can not properly be considered a common 
procedure, nor has it earned a place in the physician's laboratory. 

Significance of Chlorides. — These are normally present in the 
urine. Continued fever or serious kidney lesions cause decrease 
of chlorides. Cessation of excretion of chlorides in urine forms 



1 Only the urates are dissolved, and such quantity of the acid itself as may be 
present is so* small that the test loses none of its value. 



116 LABORATORY METHODS. 

the basis of an unfavorable prognosis in pneumonia. After the 
crisis is successfully passed, the chlorides are increased in amount. 
Estimation of Chlorides. — The following plan is simple, and at 
the same time very useful : 

1. Remove, if necessary, any albumin that may be present. 

2. Filter if necessary. 

3. Take 2 ounces. 

4. Add 1 drop of nitric acid. 

5. Add 3 drops of a 15-percent solution of silver nitrate. 

A normal amount of chlorides produces thick, curdy masses, 
whereas greatly diminished chlorides cause only a slight cloudiness. 
More delicate estimations are unnecessary for prognostic purposes. 

Significance and Detection of Diacetic Acid. — Diabetic coma is 
due to betaoxybutyric acid, but, as the presence of this is detected 
with difficulty, search is usually made for diacetic acid, a body 
closely related and usually formed simultaneously. Diacetic acid 
has been found in other conditions, and is of little diagnostic im- 
port, but its appearance late in sugar diabetes is of great interest 
in that a coma is likely to follow, and hence its identification be- 
comes of great prognostic interest. The sample must be fresh 
when tested, as this acid is very volatile. The urine must be added 
to the ferric chlorid, else nitration will be necessary in order to see 
the color reaction. The test is as follows : 

1. Into a test tube pour about 2 drams of a clear, strong solu- 
tion of ferric chlorid. 

2. Acid 1 or 2 drops of the urine. A wine-red color indicates 
the presence of diacetic acid. The reagent should not be added to 
the urine, as is generally advised. If compounds of the aromatic 
series are being exhibited, these will give a pseudotest. In such 
contingency, recall that diacetic acid is partially decomposed by 
boiling the urine just prior to the test, thus reducing the intensity 
of the reaction. 

Collecting the Sediment. — Whether gravity or the centrifuge is 
employed to separate the sediment from the liquid is immaterial, 
except perhaps that the first method requires too much time. The 
sediment may be collected from beneath the supernatant liquid by 
the following method : 

1. Use a long pipette. If a medicine dropper is employed, the 
technic may be modified to apply to its use. It is sometimes ad- 
visable to select this sediment from the bottom of a bottle, and 



THE URINE IN DISEASE. 117 

therefore a small glass tube drawn to a point should be a part of 
every physician's equipment. With the index finger pressed 
against the top, immerse this pipette so that the point comes in 
contact with the sediment. 

2. Now carefully relax the pressure on the upper end, and some 
of the sediment will be drawn into the tube. Quickly apply the 
pressure and remove the pipette from the sample. In case the 
sediment appears to be in strata, specimens from the several layers 
may be taken. 

3. On a piece of window glass (2x2 inches) drop at different 
places some of the sediment, and examine with a low-power ob- 
jective and narrowed diaphragm. In case there is any question 
concerning the recognition of certain elements, drop on a cover 
glass, slightly open the diaphragm, and use a higher magnification. 

Pus. — Separate chemical and microscopical considerations of 
pyuria are almost impossible. When its origin is in the kidney 
tissue, only a few cells are usually found, and the companionship 
of certain other elements — as casts, crystals, and epithelial cells — ■ 
may aid in determining the source. A renal pus shows a tendency 
to appear and disappear at intervals, is acid, shows no tendency 
to thread formation, and its cells are usually well preserved. 

In cystitis, if acid, differentiation may be impossible, but if alka- 
line, as is usually the case, the cells tend to be swollen or destroyed. 

In urethritis the first urine voided should contain much pus, 
while that collected in a second glass exhibits but little. It shows 
a tendency to thread formation — clap threads. 

The pus cell is usually a polymorphonuclear leukocyte, but 
lymphocytes may be present. There should be little difficulty in 
ruling out epithelial cells and unnucleated red corpuscles, and in 
case of doubt the nuclei may be brought out by adding to the drop 
of sediment a drop of 1-percent acetic acid. The pus of pyelitis is 
differentiated with difficulty. When desirable, smears and stains 
may be made for the tubercle bacillus, but the search is too often 
very discouraging. 

Casts — Etiology, Types, and Differentiation. — A complete clas- 
sification of these elements is left to the larger books. For prac- 
tical purposes, it is not necessary to draw fine distinctions — any 
type may be expected in the several kidney lesions, although certain 
forms may be more typical of an interstitial inflammation than of 
a parenchymatous nephritis. 



118 



LABORATORY METHODS. 



The recognition of tube casts should not be a difficult matter, 
but nevertheless much confusion exists in regard to this point 
among practitioners. There is no doubt, however, that one who 
has once really seen and identified these elements is never perplexed 
in the future. Some of the more common sources of error are the 
following : 

1. Mistakes in identity. The harmless epithelial cell and the 
extraneous cotton fiber are only too often mistaken for casts. 

2. Working with too much light. Many persons neglect to 
properly narrow the diaphragm in making examinations of the un- 
stained preparations of urinary sediment, as well as of other smears 
and sections which have not been treated with a dye. 

3. Attributing to the flexible and mucus-like cylindroids unde- 
served individuality. 






ir 



Fig. 2 9. —Artifacts. 




A, granular casts; B, vegetable fibers: 
lost their nuclei. 



C, epithelial cells which have 



The cast is formed in the uriniferous tubule, and the latter may, 
in fact, be considered its mold ; so that, if there is blood in these 
tubules, there will be a blood cast ; if there is pus, a purulent cylin- 
der will result, etc. The granular casts represent, in all probability, 
necrotic epithelial cells which line the uriniferous tubules ; in fact, 
certain of these casts contain large remnants of these cells or the 
cells themselves. There may be other casts representing degenera- 
tive or necrotic changes, as certain hyalin or fatty casts. When 
large numbers of bacteria are included, a so-called bacterial cast 
may result. 

In certain pathological conditions individual types may pre- 
dominate, although any or all of the others may be present. Thus, 
in an acute nephritis, search is naturally made for the epithelial, 
blood, and leukocyte casts, while the granular and hyalin forms 



THE URINE IN DISEASE. 119 

are more characteristic of chronic Bright 's disease. Epithelial 
casts are rarely found in simple interstitial types of kidney dis- 
ease. Bacterial casts indicate a very grave prognosis. In diabetes 
there are usually expected to be found, at some stage, short hyalin 
casts. 

A cast should not be confounded with a cotton or flaxen fiber, 
as there are few or no points of resemblance, which is clearly shown 
in Fig. 29. Often, though not always, these fibers lie on the cover 
slip, and the objective must be lowered before any of the urinary 
sediment comes into view. An epithelial cell in which the nucleus 
shows but faintly or not at all, and the edges of which tend to 
"roll," seems often to confuse the microscopist initiate. In case 
of doubt, a little acetic acid or a drop of slightly acidulated 






CD E 

Fig. 30.— Some typical epithelial cells from the urinary passages. A, squamous cell of 
vagina and urethra; B, caudate cells from pelvis of kidney, ureter, and bladder; C, 
cylindrical cell from the upper portion of the male urethra; D, polynuclear cell, same 
origin as tailed cell; E, two renal cells. 

methyl green may demonstrate a nucleus. Do not forget to nar- 
row the diaphragm, which admonition is emphasized because fail- 
ure to do this is one of the chief reasons why most practitioners fail 
to see casts. A real cast is, however, rarely mistaken 'for an epithe- 
lial cell or a bit of extraneous matter, as the outlines are too clearly 
cut and the morphology is too characteristic. It is therefore a 
fairly safe conclusion that where doubt exists the element in ques- 
tion does not properly belong to the pathologic nomenclature. 
Casts are demonstrated with difficulty in a urine voided several 
hours before. 

Blood Cells. — Blood- is identified microscopically by the presence 
of red cells. These, unless hemorrhage is recent, are rarely intact, 
but are swollen, and usually lack some of their pigment, at times 
even forming blood shadows (Fig. 30). 1 Casts signify renal 



1 It follows that in some urinary specimens, especially those where the examination 
has been delayed, the erythrocytes can not be identified by the microscope and the 
presence of hemoglobin must be demonstrated. For this purpose Meyer has devised 



120 



LABORATORY METHODS. 



hemorrhages. Here, as with pus, the coexistence of certain other 
elements may indicate the location of the bleeding point. 

Epithelial Cells. — A few desquamated epithelial cells may be 
found in a sample of normal urine, and large numbers of these 
elements point to pathological processes. Unfortunately the shape 
of the cell rarely aids in diagnostics, as the various types are so 
widely distributed. Ordinarily the characteristic cell of the va- 
gina or urethra is referred to as a flat cell (Fig. 30). Vesicular 
denudations, as well as those from the ureter and kidney pelvis, 
usually include characteristic star-shaped and "tailed" cells. 
Very small round cells, with spherical nuclei, are most likely to come 
from the uriniferous tubules. 

Chemical Sediments. — These are of less diagnostic significance 
than the organic elements. The following is a brief classification 
of the more common crystals : 




Fig. 31. — Comparison of the more usual forms of the common crystals met in urinary sedi- 
ments. A, uric acid; B, ammonium urate; C, calcium oxalate; D, triple phosphate; 
E, calcium carbonate. 

1. Uric Acid and Urates (Brown Color). (Fig 31.) Soluble 
only in warm solutions. Typical uric acid crystals are of a rhom- 
boid shape, and those of ammonium urate have an appearance not 
unlike that of the common cockle-bur. There are many variations 
from these characteristic forms, but certain points of resemblance 
are found in the less frequently observed types. Occur in normal 
urine, but are increased in gout. 

2. Calcium Oxalate. Show typical envelope shapes. Soluble in 
hydrochloric acid and insoluble in acetic acid. Of little patho- 



an excellent test. Prepare the reagent by heating 4 grams of pure phenolphthalein, 20 
grams of potassium hydrate, about 15 grains of zinc dust, and 200 cc. of distilled 
water. Prolong the heating until all the pink color disappears, and then filter and 
bottle. Having obtained a good solution, which may be kept a long time, the test 
itself is simple. Add a few drops of this reagent and a drop of peroxid to a few cubio 
centimeters of the urine. A pink coloration indicates the presence of hemoglobin. 
Make a control test with a normal urine. 



THE URINE IN DISEASE. 121 

logical significance. When present in large amounts, these crystals 
may irritate the ureters and give rise to symptoms similar to those 
noted in renal calculus. Such a condition has been termed pain- 
ful oxaluria — oxaluria dolorosa. 

3. Triple Phosphate. Heat causes precipitation rather than the 
solution noted with urates. The coffin-lid shape is typical. Oc- 
curs in ammoniacal fermentation, whether in a diseased bladder or 
in the urinal. 

4. Calcium Carbonate. Occurs as small transparent spheres, 
which give off gas (carbon dioxid) if acid is added. Occur under 
the same conditions as the triple phosphate. 

Bacteria in General. — A few bacterial cells may be present in a 
normal urine freshly voided into a sterile receptacle. Urine, on 
standing, soon becomes contaminated by microorganisms of the 
air, and "spoiling" occurs. In a majority of cases this fermenta- 
tion is alkaline in character (ammoniacal), and such a urine 
abounds in germs of many varieties. 

Parasites. — The tubercle bacillus may, with difficulty, be identi- 
fied in cases of tuberculosis of the genital tract. See Searching for 
Germs (page 48) for the detection of the gonococcus. The micro- 
organisms of many system affections, as typhoid, may occur in the 
urine, but are rarely considered from a diagnostic standpoint. The 
colon bacillus is usually blamed for an acid cystitis, whereas a 
bladder affection alkaline in character is often due to the bacillus 
proteus vulgaris. Multitudinous varieties of bacteria abound in 
ammoniacal fermentation of the urinal. Smegma bacilli, yeasts, 
molds, amebae, and the eggs and bodies of certain animal parasites 
have been identified, but, with the exception of the first of these, 
are very rare. It follows, therefore, that a diagnosis of colon 
cystitis on the appearance of rods in a sample urine is as imprac- 
tical as the diagnosis of consumption on the presence of a cough. 
Strange to say, many of these impractical procedures are often 
conducted by men who acknowledge that they are unable to identify 
a urinary cast or differentiate between albumin and urates. 

Extraneous Matter. — The various vegetable fibers and starches 
which may be found in any microscopical preparation have been 
previously considered (page 30), and should be easily recognized. 
They are introduced accidentally as contaminations from the hands, 
towel, or other sources not easily determined. 

Ammoniacal Fermentation. — AYhile reference has been made to 



122 LABORATORY METHODS. 

this condition (page 108), a few special points may be emphasized. 
A "spoiled" urine may be found either in a cystitis or in a stink- 
ing urinal. In the former case it is most likely to be due to the 
bacillus proteus vulgaris, but when voided may be caused by many 
types of microorganisms. Under the alkaline cystitis (page 122) 
the findings of this condition have been listed, and they differ very 
little from those here noted as the fermentation in the urinal: 

1. Pungent odor, due not only to ammonia gas, but to certain 
other decomposition products. 

2. Marked alkaline reaction. 

3. Presence of many bacteria. 

4. Dirty white or gray sediment, which shows under the micro- 
scope the coffin-lid crystals of the triple phosphates and the spher- 
ical carbonates. 

Urine of Specific Urethritis. — Contains clap threads, pus cells 
containing gonococci, and rarely blood. 

Urine of Acid Cystitis. — Acid reaction. Slight fecal odor. Con- 
tains pus, red blood cells, and epithelium. Caused commonly by 
the colon bacillus. 

Urine of Alkaline Cystitis. — Marked alkaline reaction. Foul 
odor of ammonia and putrefactive gases. Contains triple phos- 
phates, pus, epithelium, and rarely blood. Caused commonly by 
bacillus proteus vulgaris. 

Urine of Foudroyant Cystitis. — Characterized by much blood 
and pus. Caused by streptococci and other virulent germs. 

Urine of Pyelitis. — Rarely shows any features different from 
those of acid cystitis. 

Urine of Renal Calculus. — Acid reaction. Contains blood and 
often bits of urate or oxalate gravels. 1 

Urine of Vesical Calculus. — This presents a picture of a severe 
cystitis, with blood and possibly phosphatic concretions. 

Urine of Renal Hyperemia. — Small amounts of albumin, but 
few or no casts, with little or no blood. In the passive hyperemia 
of mitral insufficiency the urine is acid, contains many urates, and 
has a high specific gravity. 

Urine of Uremia. — Many casts of all types, much albumin, and 
decreased urea. 

Urine of Parenchymatous Nephritis. — Small amount of urine, 

1 The presence of oxalate crystals in the urine is not sufficient proof that a calculus 
is lodged in the renal pelvis or ureter. That these crystals may cause distressing 
symptoms resembling those of renal calculus appears to be unquestioned 



THE URINE IN DISEASE. 123 

containing much blood and albumin, and of a high specific gravity. 
Contains, besides free blood, blood casts and many epithelial 
casts. 

Urine of Interstitial Nephritis. — This is the chronic Bright 's 
disease. It is characterized by much urine, containing no blood 
and only traces of albumin, and shows a low specific gravity. Hya- 
lin and granular casts are usually present, but the others are 
rarely found. Cellular elements are generally few in number. 

Urine of Diabetes Mellitus. — Much urine, usually of a high 
specific gravity. Smells sweet and has but little color. Glucose 
and the acetone bodies are present. Traces of albumin and the 
short hyalin casts are not uncommon. 

Urine of Diabetes Insipidus. — Much urine of a low specific grav- 
ity, but shows no sugar, acetone bodies, albumin, etc. 

Urine of Autointoxication. — Variable but usually small amount 
of urine of a high acidity, sometimes showing traces of albumin 
and hyalin casts but invariably contains large quantities of in- 
dican or indolacetic acid. Tests for these as well as a bedside 
method of estimating the acidity, may be found in the appendix. 
Some authorities term this condition acidemia or copremia. 

Acidosis Versus Acidemia. — Harrower and other writers have 
pointed out these differences. A brief classification of all that 
pertains to the acids of the urine is here given : 

1. Normal Acidity. To certain phosphates is due the normal 
urinary acidity. 

2. Uric Acid Diathesis. It has yet to be proven that excess of 
uric acid in the urine increases the acidity of that excretion. 

3. Acidosis. When the urine of sugar diabetes contains the 
acetone bodies. 

4. Acidemia. A condition observed in autointoxication, and un- 
doubtedly caused by certain acids containing sulphur, by indola- 
cetic acid, and by other products formed in abnormal intestinal 
putrefaction. 

Tuberculosis of the Urinary Tract. — The tubercle bacillus is not 
easily found, which is especially the case if much blood is present, 
and it is well to centrifuge before making the spreads. The 
smegma bacillus, though acid fast, shows no tendency to occur in 
clumps. If any doubt exists, it may be advisable to wash the 
stained preparation in an absolute alcoholic solution of 1-percent 



124 LABORATORY METHODS. 

hydrochloric acid for ten minutes, which will decolorize the smegma 
bacillus, but the tubercle germs will remain brightly stained. 

Iodin Tests in Malingerers. — Under certain conditions a physi- 
cian may desire to know whether his patient is taking the medicine 
as prescribed or dispensed. By adding a few grains of potassium 
iodid and collecting a sample of the urine, that drug may be 
easily detected as follows: To the urine add a little starch solu- 
tion, and then add 1 or 2 drops of chlorin water. A blue color 
indicates the presence of the iodid. 

Tampering with a Urine. — The authors have observed several 
instances of this kind. A quack doctor taught his patients to test 
for albumin and sugar, and then prescribed at intervals the oil of 
copaiba. One woman, insistent that she had diabetes, was finally 
informed that her urine contained large quantities of cane sugar. 
The addition of glucose to a voided urine is quite common, and 
Osier records one case where it was introduced into the bladder. 
Brick dust or iron rust may be added to give the appearance of 
urates or blood, and ammonia may be added, which does not, how- 
ever, result in the formation of the typical coffin-lid crystals of 
triple phosphate, but in certain star-like crystals. A candidate for 
a life insurance policy may bring a bottle of normal urine to the 
office, whereas a freshly voided sample from the same person would 
show sugar, albumin, or casts. The physician must be on the look- 
out for deception. 

Less Frequently Applied Procedures. — Some tests have no di- 
agnostic or prognostic worth, and others of great value are rarely 
or never attempted by any save experts. Both of these classes have 
been combined and listed as follows : 

1. Temperature corrections for specific gravity. 

2. Detection and estimation of certain salts and other constitu- 
ents of the urine. 

3. Several tests for albumin and its quantitative estimation. 

4. Certain bacteriological examinations, etc. , 
Value and Limitation of a Practical Urinalysis. — The value of 

the tests described depends on their proper selection and applica- 
tion, and, while their importance is too often underestimated, the 
deductions drawn from this source alone are of little value. The 
urinalysis is only another link in that chain by which a safe diagno- 
sis is determined. 



CHAPTER XI. 



MILK AND ITS HOME MODIFICATIONS. 
Apparatus. — 



1. Hydrochloric acid, dilute. 

2. Sulphuric acid, commercial, slight- 

ly diluted. 

3. Sulphuric acid, C. P., concentrated. 

4. Methyl alcohol. 

5. Fusel oil. 

6. Ammonia water. 



10. 
11. 



Centrifuge and special tube. 
Microscope and accessories. 
Phenolphthalein, 1-percent alco- 
holic solution. 
Pipette or medicine dropper. 
Tincture turmeric. 



12. Urinometer. 



The methods given here are those which are used successfully 
by the leading pediatrists of the country. So far as practical pur- 
poses are concerned, the formnlas for the home modifications are 
ideal in that the mixing may be done by the housewife. These 
tests and estimations are exceedingly simple, but the lack of proper 
references renders it necessary to include in this chapter more de- 
tails than might be desired. Moffitt, in the Journal of the American 
Medical Association, has ventured to criticise the absolute accuracy 
of the formulas, but his observations are at variance with those of 
the authors, who have noted that control analyses of samples after 
modification seem to be of surprising accuracy. It is the opinion 
of many authorities that it is unnecessary for these to be abso- 
lutely accurate. For the benefit of those not so well informed on 
the subject the authors have deemed it advisable to enter into detail. 

Substitutes for Mother's Milk. — The summer months is the time 
when the health of the baby is generally affected, and when its 
food must be changed it may occur to the physician that the little 
patient needs modified milk, but he makes no effort to put his 
opinion into practice. The home modification of milk and the 
principles of infant feeding are so little understood by the profes- 
sion that any light 'thrown on the subject must be of more than 
passing interest. Observations in a number of cases will convince 



References. — Lowenburg: Journal of American Medical Association, vol. LV, No. 7, 
page 565; Kerley : Diseases of Children; Koplick: Pediatrics; Holt: Diseases of Infancy. 

125 



126 IjAboratort methods. 

the physician that there is a science in feeding the baby which is 
denied the maternal milk. 

The earliest work in this matter was done by T. M. Rotch and 
G. E. Gordon. The work of the Walker-Gordon laboratories is to 
be commended, but hardly comes within the scope of this book, as 
they are rarely accessible to the general practitioner. They furnish 
the milk direct on prescription, acting in the same capacity as the 
pharmacist, and merely see that the patient gets exactly what is 
ordered. A physician who is ignorant of the principles of infant 
feeding can place no blame on these laboratories for poor results. 
The absence of these facilities should not, however, render helpless 
the man outside of the larger cities, as an energetic physician, an 
intelligent mother, and a conscientious milkman can accomplish 
just as good results as the best equipped metropolitan laboratory. 

Maternal nursing is most to be desired. The woman who refuses 
to nurse her child deserves censure, but the physician is often an 
unsuccessful social reformer. Not only is the mother's milk more 
acceptable to the infant, but nursing the child aids the uterus to 
contract and involute properly, and especially among the poor and 
ignorant should maternal nursing be encouraged. Severe post- 
partum hemorrhage, acute infections, tuberculosis, and certain 
chronic nervous diseases, as epilepsy, are contraindications. Ma- 
ternal syphilis, where the child seems to be healthy, should be con- 
sidered a contraindication, although the healthy parent may nurse 
with safety her syphilitic babe. 

The best substitute for the milk of the mother is that of some 
other woman, but in America there are no slaves or peasants, and 
a healthy wet nurse is either a luxury or an impossibility. Hid- 
den disease is always a menace to both parties. Mixed feedings 
may be deemed advisable where the milk of the mother is good, but 
small in amount. 

Of all artificial foods, modified milk is accepted by scientific 
men as ideal. Its real merits are borne out by clinical experience, 
and the results of its therapeutic efficacy may be seen in a hospital 
ward furnished with scientifically prepared modified milk from 
the adjoining milk laboratory. The baby's organs of nutrition, 
however, form a machine doubtless more complex than that of the 
adult, and no physician should become so infatuated with this 
method of feeding that he would see in it perfection, as there is 
still much to learn about feeding the baby. Because of disappoint- 



MILK AND ITS HOME MODIFICATIONS. 127 

ments to those who were at first too optimistic and the ignorance of 
the profession as to the simplicity of the home modification, the 
various proprietary infant food concerns exist. The products of 
these concerns may have done some good, but unfavorable results 
do occur from their use, and experience shows that these unfavor- 
able results are the rule. In the first place, they are stock pre- 
scriptions, and in their preparation the condition of the patient 
is not and can not be taken into consideration. Any intelligent 
physician will regard this "panacea" from about the same view- 
point that he would look on any other cure-all. A beautiful book- 
let may give directions how to mix it with cow's milk, but only the 
latter is needed, as it is cheaper, safer, and better for all babies, 
sick or well, and its real worth is shown by the attempts to imitate 
it. These statements are not made in an unfriendly spirit, but cer- 
tain comparisons are presented for the guidance of the general 
practitioner. 

Proprietary Foods Versus Modified Milk. — There have been 
cases in which the proprietary foods have been prescribed where 
they either fortunately suited the condition or were selected by a 
physician who understood the principles of feeding and knew the 
composition of the food, and who applied this knowledge to the 
particular infant. Several brands may have been cast aside as 
worthless in an exhaustive experiment, and one found to fit the 
case just in time to avoid fatal results. The picture of the healthy 
infant is shown in the advertisements, but the others who developed 
rickets, vomited up their food, or died are not portrayed. 

The following are the advantages and disadvantages of the pro- 
prietary foods, as well as those of cow's milk scientifically modi- 
fied: 

Advantages of Proprietary . Foods. 

1. Convenient to use. 

2. Suit some babies under some conditions. 

3. Fairly good substitutes, when properly modified, for cow's 
milk when the latter can not be obtained in a pure state. 

Disadvantages of Proprietary Foods. 

1. Expensive, and usually unnecessarily so. 

2. Not ideal substitutes from a scientific standpoint, as the pro- 
teids, fats, and carbohydrates are not in correct proportion even 
for the healthy baby. 

3. Not ideal substitutes from a clinical standpoint. Unless the 



128 LABORATORY METHODS. 

food happens to suit the case, there results scorbutus, rachitis, 
marasmus, or even acute inanition. 

4. They present an attempt to fit the patient to the cure. 

5. The proteids and sugars of the cereal foods are entirely dif- 
ferent from those of the animal milks, and, although their amounts 
may be modified, an alteration of their composition is impossible. 
Because "proteids are proteids" is no reason why bad proteids 
should be substituted for good proteids. All sugars are sweet, but 
the diabetic may safely take levulose when glucose may be poison- 
ous to him. Sugar of milk is absent in many of these foods. 

6. Menace infantile life because they are offered to the laity, 
and the helpless mite is intrusted to the untrained. They are pre- 
scribed according to bottle directions, and the parents, having the 
physician's recommendation, are under the impression that science 
can do no more than they in saving the baby. 

7. Menace infantile life because in many cases the manufacturers 
presume on the ignorance or indifference of many of the profes- 
sion, so that the real dangers are hidden. 

8. Given straight or modified, no artificial food contains those 
milk enzymes known to be so necessary to the infant, unless it also 
contains that which is absolutely necessary — the fresh milk of some 
mammal. 

Advantages of Modified Cow's Milk. 

1. Approaches nearest to mother's milk, and may be further 
modified to suit the infant vrhen even mother's milk, if obtainable, 
would fail. 

Disadvantages of Modified Cow's Milk. 

1. AVidespread belief that the process is very difficult. 

2. Ignorance of the profession not only of the process of modi- 
fication, but of the principles of infant feeding. 

3. Lack of laboratory facilities. 

4. Indifference even on the part of recent graduates, who should 
know how to feed babies. 

Working in the Dark. — Cow's milk is crudely modified by some 
physicians by decreasing all the ingredients simultaneously by 
diluting the milk in toto ; then, if it is desired to bring up the fat 
content, cream is added, or, if the sugar is to be increased, sac- 
charum lactose may be added. It is seldom necessary to increase 
the proteids, as there are enough or too much in cow's milk. At 
best the system is one of working in the dark, as cow's milk 



MILK AND ITS HOME MODIFICATIONS. 129 

varies greatly. The reason that even rickets may result may be 
seen in the following table, in which the proteids of the cow 's milk, 
by dilution with equal parts of water, have been brought down to 
coincide with those of woman's milk, but there is a great variation 
in the fats. 

Proteids. Fats. Carbohydrates. 

Cow's milk 3 percent. 4 percent. 5 percent. 

Cow's milk diluted. .1.5 percent. 2 percent. 2.5 percent. 

Woman's milk 1 percent. 4 percent. 7 percent. 

There must, of course, be a good milk supply, and to assure this 
a testing of milk for dilution, cleanliness, etc., appears necessary, 
though a conscientious milkman may materially aid the physician 
in this matter. The family of the young patient can also aid in 
perfecting a suitable modified milk, but the physician should thor- 
oughly understand the principles of infant feeding. 

The Laboratory. — This is a secondary matter, but important. 
Each physician may have his own work-room, fitting it up for less 
than he pays for paregoric and infants' anodynes; or several physi- 
cians may have a laboratory in common. The worker needs a bot- 
tle of sulphuric acid and a few other reagents, and, if he possesses 
a centrifuge, he may obtain a fat testing tube at small expense. 
For convenience of description, we shall consider the laboratory 
work and the prescribing as being done by two different men. The 
actual mixing of the baby's milk is done every day by the family 
according to directions from the laboratory man, who is in turn 
instructed by the physician in charge of the case. 

Writing the Prescription. — The physician may be his own 
analyst, but, in case another man does the work, his relations to the 
latter are as follows : 

1. He should devise a prescription to suit the case, and, if he is 
not sure as to the one indicated, should study up the subject. 

2. He should see that a sample (one quart) of the milk reaches 
the analyst. In many cases the milk dealer should not be aware, 
for baby's sake, that this examination is to be made. 

3. The prescription should state the amount of milk required for 
the infant every twenty-four hours. 

At this point the responsibility of the prescriber becomes that of 
the laboratory worker, but it is advisable to enlarge on these three 
points before we study the work of the milk laboratory. 



130 LABORATORY METHODS. 

Let us first consider the devising and writing of the prescription. 
Many of the poor results from using modified milk are due to igno- 
rance in this matter. A physician should not merely' write 

R Modified Milk 5 v 

Sig., etc. 

As has been stated, a knowledge of the principles of infant feed- 
ing is necessary, and, if this is lacking, it may be obtained in a 
short review of a book on pediatrics. The authors do not wish to 
overstep the scope of this volume, but emphasizing a few principles 
will not be out of place. 

Some Principles in Baby Feeding. — There are two numbers 
which every physician should bear in mind — viz., 453 and 471. 
Although the makeup of woman's and cow's milks vary, for prac- 
tical purposes the following arrangement will answer: 

Fats. Carbohydrates. Proteids. 

Cow's milk 4 5 3 (453) 

Woman's milk 4 7 1 (471) 

First let us consider the healthy baby where maternal milk can 
not be obtained. Many of the bad effects of unmodified cow's milk 
in the healthy infant is due to the high proteid content of that 
secretion. The carbohydrates, although a lesser factor in these 
considerations, should also be changed in order that mother's milk 
may be approximated as nearly as possible. For that reason a 471 
milk may be tried, but experience shows that when beginning a 
modified feeding it is often best to reduce also the fats, and a popu- 
lar trial mixture has resulted in the 261 combination. If the lat- 
ter does not agree with the baby, it may be modified, and symptoms 
should be the guide for modifications. Gastric symptoms indicate 
fat indigestion, and this substance must be increased or lowered 
accordingly — generally lowered. When intestinal symptoms pre- 
dominate — either diarrhea or costiveness — the proteids are at fault. 
Early symptoms of rickets indicate that the fats must be increased 
at once. Such symptoms as sweating of the head, restlessness at 
night, constipation, beading of the ribs, and craniotabes are im- 
portant, but easily overlooked. Stool analyses rarely hold that 
value observed where the infant is sick, and the apparently patho- 
logical stools which may be passed by the healthy infant are sur- 
prising. 



MILK AND ITS HOME MODIFICATIONS. 131 

For the sick baby we must use those principles indicated in the 
text books, and we must not forget our drugs — castor oil and 
peppermint water. Contrasted with the healthy infant, the ap- 
pearance of the stools should be noted, as this is of great im- 
portance. (See Every-Day Stool Tests, page 154.) An entire 
change of the prescription or its abolition may be necessary. 

When the baby becomes older, or even with the sick young baby, 
it may be advisable to add certain substances found neither in 
the milk of the cow nor that of the human, but which clinical ex- 
perience has demonstrated to be useful should certain indications 
arise. These have but little to do with this subject, and are really 
not foods, but tend toward drug medication. They have been used 
in the city laboratories for the convenience of physicians, and 
under this head come the syrup of lime, peptonization, barley 
water, whey, and buttermilk feedings. They should be used under 
the direction of the physician in charge, but, if dispensed, should 
not, like all other prescriptions, be refilled without his sanction. 

Sample for Analysis. — This investigation is usually necessary 
for many reasons. In the first place, all milks differ in fat con- 
tent. The variations in the other constituents are less marked, 
and usually need not be considered. The amount of fat in a given 
milk varies according to the breed of the cows, the time of the year, 
grazing conditions, first milk or top milk, whether the milk comes 
from a single cow or a herd, etc. Even if the composition of all 
milks were constant, the ignorance of many dealers renders it ad- 
visable to always be assured of their purity. Dr. Woods Hutchin- 
son says that one teaspoonful of milk may contain more inhabitants 
than the city of New York. If these were all lactic acid bacilli, 
no harm would result, but there is no natural selection. Dr. 
Hutchinson states that 90 percent of the injurious effects of milk 
is due to the germs of plain, common dirt, and this comes not only 
from the barnyard, but from unwashed milk cans and other avoid- 
able sources. Asepsis is no less important here than elsewhere, 
and sterilization, except for long journeys, is rarely necessary and 
even harmful in many cases. 

Tampering with Baby's Food. — We are too often shocked to 
learn that the apparently honest milkman is disposed to tamper 
with his milk, and evidently infanticide is justifiable in his eyes. 
He will add water to increase his profits, but, worse, he will add 
some poison to inhibit the lactic acid bacillus, unmindful of the 



132 LABORATORY METHODS. 

other varieties which nevertheless proceed to multiply. He re- 
moves the danger sign — the milk does not "sour," and it is impos- 
sible to determine whether it is fresh or spoiled. The milkman is 
saved the expense and trouble of ice, but the consumer is subject 
to his poisons. Analysis of the milk guards against these dangers, 
but it is advisable to select a milk with an apparently good cream 
content and one apparently clean. 

Sterilization is never advisable, and pasteurization is permissible 
only under unusual conditions. Some of the germs may be killed, 
but their poisons prevail. The sugar is changed to caramel, the 
taste of the milk is altered, there may be some coagulation of the 
proteids, the enzymes are destroyed or altered, and constipation 
usually results. Scurvy is caused no less by sterilized milk than 
by the proprietary foods. Fresh milk, ice, and cleanliness are 
points which need emphasis. Mechanical filtration through cotton 
is never a safe procedure. It is best, when possible, to arrange 
with some honest dealer for the milk of some certain cow or herd, 
and to insist on cleanliness, being specific as to directions and, if 
necessary, paying more for this milk; but even then certain analy- 
ses may be considered advisable. 

Amount Used Daily. — The laboratory man must know the 
probable amount of milk necessary for the day's feeding, which is 
usually determined with ease. In case the babe has never taken 
artificial food, an estimate may be made and the formulas subse- 
quently modified according as to whether the amount is too large 
or not sufficient. The amount will vary with the age of the child, 
the number of feedings daily, state of health, etc. 

Duties of the Physician. — If the physician does not do the actual 
work, he will write the prescription to suit the case, and see that 
a sample of the milk reaches the laboratory. 

Duties of the Analyst. — Each physician, as has been stated, may 
do his own work, or at least one physician in each community 
should be prepared to make the tests and estimations. The ex- 
pense of equipment is small, and the technic as described in this 
work is exceedingly simple. 

1. General appearance of the milk as to color, odor, amount of 
cream, etc. 

2. Specific gravity. 

3. Reaction. 



MILK AND ITS HOME MODIFICATIONS. 133 

4. Microscopy. 

5. Tests for chemical purity. (When searching for germs, refer- 
ence may be made to Searching for Germs, page 37.) 

6. Quantitative estimation of fat content. 

7. Modification, which consists of devising the amounts of cream, 
milk, water, and milk sugar to be used, bearing in mind the follow- 
ing factors : 

(a) Composition of the sample as determined by the tests. 

(b) The physician's prescription. 

(c) Amount to be made up daily. 

(d) Certain stock formulas which experience has shown will 
give mixtures that, when subjected to analysis, show the desired 
composition. 

A study of the forms below, being similar to those used in hos- 
pitals, will give an idea concerning the meaning of the above ar- 
rangement. The first is a prescription written for a healthy baby 
which is denied the maternal milk, followed by the report of 
directions, etc. 

R Fats 2 percent. 

Carbohydrates 6 percent. 

Proteids 1 percent. 

Amount daily 20 fluidounees. 

Report. Milk normal, the cream containing 16 percent fat. 
Modification. As follows : 

Cream 1% fluidounees. 

Lactose 1 ounce. 

Milk 3y 3 fluidounees. 

Water 15 fluidounees. 

The above milk is mixed by the family. It is made up from 
cream, milk, and water to the 20 fluidounees and then the sugar is 
added. It will be remembered that when a solid dissolves in a 
liquid, the bulk is not perceptibly increased. 

Analysis of Cow 's Milk. — Color. This should vary from a white 
to a bluish tint, according to the amount of cream. Certain ab- 
normal colors may be due to bacteria. The color of the collustrum 
is yellow, and needs no comment. The following arrangement will 
aid in the classification of such milks : 



134 LABORATORY METHODS. 

Color. Etiological agent. 

Blue Bacillus cyanogenus. 

Violet Bacillus violaceus. 

Pink Bacillus prodigiosus. 

Red Bacillus lactis erythrogenes. 

Yellow Bacillus synxanthus. 

Gelatinous color and consistency Bacillus lactis viscosus. 

Blood may color milk from a bright-red to a dark-brown. 

Odor. Abnormal odors are rarely due to added preservatives, 
as these occur in very small amounts. The gelatinous milks may 
possess a very offensive odor, and, strange to say, the most dis- 
gusting of these cause no symptoms when ingested, whereas an 
apparently perfect milk may teem with death-dealing microorgan- 
isms or contain formaldehyd or borax in very small quantity. Cer- 
tain vegetables, as turnips, cause a distinct and characteristic odor. 
A bitter milk may be explained by the presence of certain bacteria, 
purposely added poisons, etc. Milk absorbs odors, especially when 
stored in ice chests with melons, cucumbers, canned salmon, etc. 

Amount of Cream. In a regular pint bottle which has stood over 
night the upper cream layer should by linear measure make up 
about one-third of the entire amount. 

Specific Gravity. This may be taken with an ordinary urinome- 
ter and should average 1.030. In case it is much more, there is 
sufficient reason to suspect that cream has been removed. If it is 
considerably less, the milk has in all probability been watered. 

Reaction. Milk is acid when tested with phenolphthalein solu- 
tion. It is practically never alkaline unless putrefactive changes 
are present or chemicals have been added. Acidity is increased as 
the milk sours. 

Microscopy} If the cow is healthy, only a few leukocytes and 
epithelial cells may be present in her milk. Leukocytes in con- 
siderable number point to infection. 

Detection of Formaldehyd. To 4 drams of milk in a test tube 
add 1 dram of commercial sulphuric acid. Do not mix, but permit 
the acid to form a layer below the milk. A violet ring at their 
junction is proof that formaldehyd is present. This test is ex- 
ceedingly simple and delicate, and but two precautions are neces- 
sary : 



1 Experts are able to obtain merely from a microscopical examination considerable 
information as to the probable percentage of fat. A granular tendency of the droplets, 
or when these are much too large, points to a paucity of fat. 



MILK AND ITS HOME MODIFICATIONS. 



135 



1. The reaction depends on the presence of iron contained in the 
commercial sulphuric acid ; hence, if pure sulphuric acid is used, 
two drops of ferric chlorid should be added to it. 

2. A pseudo-reaction may occur if acid is concentrated, or the 
charring of the milk may obscure the true reaction ; hence an acid 
of about 1.700 specific gravity — i. e., slightly diluted — should be 
employed. 

Detection of Borax and Boric Acid. Mix 
in an evaporating dish 1 dram of the milk with 
1 dram of the fresh tincture of turmeric and 
heat, evaporating slowly to dryness. Add 2 
or three drops of hydrochloric acid (dilute) 
to the residue and evaporate to dryness once 
more. Pink or red colorations are positive in- 
dications, and a drop of ammonia water should 
change either to a green. This test takes some 
time, and is rarely necessary as a routine pro- 
cedure. A milk which shows no formaldehyd 
and which refuses to sour is likely to contain 
borax or boric acid. The chief source of er- 
ror is too rapid evaporation by heat from the 
free flame. A good substitute for the water 
bath is to place the evaporating dish on the flat 
lid of a kettle of hot water. 

Detection of Sodium Bicarbonate. Its prob- 
able presence may be determined by adding a 
drop of any inorganic acid — for example, 
nitric acid. In case soda is present, gas will 
he given off and the surface of the milk will 
froth. In case other alkalies are added, a 
drop of any chemical indicator will show at 
cnce the nature of the deception. The value 
cf these tests would be limited in a decom- 
posed milk. ' 

While it is true that tests for chemical preservatives need not 
be made in all cases, yet positive reactions are often found where 
they might be least expected. Such poisons as have been men- 
tioned are not usually added to milk during the colder months. 

Quantitative Estimation of Fat. This is necessary with all 
creams, except perhaps those in which previous test has shown a 



;c.cj 

I 



32 — Tube for fat 
estimations which 
may be used with an 
ordinary centrifuge. 



136 LABORATORY METHODS. 

definite percentage of fats. For a few cents a special tube (Fig. 
32) may be purchased which may be used with the ordinary centri- 
fuge. A mixture of 1 part of cream and 4 parts of water is added 
with a pipette to the 5 cc. mark. Add 1 drop of the Leffman-Beam 
solution, made up as follows : 

R Hydrochloric acid 50 drops. 

Methyl alcohol 13 drops. 

Fusel oil 37 drops. 

Mix by shaking, and then add concentrated C. P. sulphuric acid 
drop by drop, shaking constantly or rotating tube until the zero 
mark is reached. Centrifugalize five minutes and then read per- 
centage of fats directly from scale. Multiply by five, which gives 
the percentage of fats in the sample of cream. 

Formulas. — These are stock formulas, and their use has been 
previously considered (page 133). The following letter symbols 
and formulas are recommended for the home modification of milk: 

Q — Quantity desired for twenty- four hours' feeding. 

F — Desired fat percentage in modified milk. 

S — Desired sugar percentage in modified milk. 

P — Desired proteid percentage in modified milk. 

FF — Percentage of fat found in the cream by analysis. 

C — Amount of cream to be used in the prescription. 

M — Amount of milk to be used in the prescription. 

W — Amount of water to be used in the prescription. 

SS — Amount of milk sugar to be used in the prescription. 

Formulas for Milk Modifications. — 

c = -f F 4r 4 x(F - p) 

M= . .— C 

4 

ss= «S-P 1 xQ 
100 

W = Q— (M + C) 

Example. Taking the above formula as a basis, substitute for 
it a 20-ounce mixture that is to contain 2 percent fats, 6 percent 



MILK AND ITS HOME MODIFICATIONS. 137 

carbohydrates, and 1 percent proteids, and where the cream has 
shown 16 percent fat. 

20 

C = X ( 2 — !) = 1% fluidounces. 

16 — 4 

20X1 
M = — 1% = 3y 3 fluidounces. 

(6 — 1) X20 

SS == = 1 ounce. 

100 

W = 20 — (3y 3 + 1%) =15 fluidounces. 

It will be noted that "Q" is made np by the addition of the 
computed quantities of cream, milk, and water respectively. The 
milk sugar dissolves without increasing the bulk to any extent. 

Less Frequently Applied Procedures. — These include mainly 
the bacteriological tests. Dr. Hutchinson's Manhattan in a spoon 
consists of as many nationalities as does the real Ghetto, from 
which it may be seen how impossible it would be for the general 
practitioner to isolate and study this population. 

Analysis of Maternal Milk. — A search for a pathological leuko- 
cytosis or an estimation of fat may be seldom attempted by the 
practitioner, but, with these possible exceptions, little practical 
information is to be gained from a study of this secretion. It 
should be understood that some of the germs of an acute infection 
may pass into the milk, and that a laboratory examination will not 
be necessary to condemn it. 

The questions of how to overcome the difficulties that may be 
met and the value and limitations of certain procedures have been 
previously considered in descriptions of the technic. 



CHAPTER XII. 

SOME SIMPLE WATER ANALYSES. 

Apparatus. — This is listed preceding each test, and necessarily 
depends on the selection of the tests made by the analyst. 

These tests, when compared with those procedures usually de- 
scribed, are simple. The recommendations made to the general 
practitioner may be criticised, but a careful study of the matter 
will show that the analysis by the chemist at a distance is not in 
every case perfectly satisfactory. In order to avoid any unfair 
interpretation of the value of these investigations, the authors ask 
that a careful study be made of this chapter. In all tests, only 
the water used for drinking purposes is considered. 

Scientific Versus Practical Analyses. — A practical examination 
of drinking water is in all cases useful and in some instances in- 
dispensable. A full and scientific investigation will reveal certain 
facts that are as yet of little or no real value, and do not receive 
consideration in this book. It follows, therefore, that any criti- 
cism of the methods that are recommended can not properly be 
offered by individuals interested commercially in the "perfected 
water laboratory." It is not the intention to recommend, without 
limitation, every investigation mentioned in this book, for in this 
matter, as in other lines of work, a certain amount of practice 
is necessary. It may be advisable, in turning over the practical 
examination of drinking waters to the physician, to state two 
propositions and then to attempt their proof : 

1. The physician may be able, after a little practice, to condemn 
a drinking water quite as quickly as an expert. 

2. No chemist or bacteriologist can, from the examination of a 
water sample, recommend it for drinking purposes, and this ap- 
plies to the expert as well as to the practitioner. That a water is 
probably safe can not be determined any quicker by the extended 
scientific examination than by carrying out the work as here indi- 



References. — Jordon : General Bacteriology; Prescott and Wilson: Elements of Water 
Bacteriology ; Harrington : Practical Hygiene. Very few books issued prior to the 
past two years bring this subject down to date. 

138 



SOME SIMPLE WATER ANALYSES. 139 

cated. It should be remembered, however, that in this chapter is 
considered only a suspected water, and not the larger sanitary 
questions — not, for example, the typhoid epidemics, where milk or 
flies may be the carriers, and where expert assistance is always de- 
manded. 

The first proposition may be proven by the actual selection and 
description of the tests, their limitations, etc., together with a thor- 
ough study of these points by the physician and actual practice 
with samples. 

The second proposition may seem more startling, and demands 
discussion at once. Unfortunately, for analytical purposes, the 
nature of the bacterial contamination of water is of much more 
importance than its actual amount. A water which yesterday con- 
tained the germs of typhoid may show none today, and these same 
germs may be found tomorrow or even immediately after the 
samples were taken. The water may be sparkling, clear, and with- 
out odor, and a careful chemical examination may reveal the 
minimum amounts of salts resulting from organic decomposition. 
A thorough bacteriological examination may neither show evi- 
dences of sewage contamination nor may it identify specific patho- 
genic microorganisms. The fact, therefore, remains that no 
laboratory analysis can at present recommend for drinking pur- 
poses any certain water. 

Analysis of Commercial Waters. — Purgative waters — those rec- 
ommended for ' ' kidney disease, " " rheumatism of the young man, ' ' 
"health waters," etc. — are not considered in these tests. They 
contain large amounts of mineral water derived from deep-seated 
natural deposits, and in most of them the saline purgatives abound. 

Detection of Poisonous Chemicals. — These are rarely found in 
drinking waters. Some western rivers have been said to contain 
in solution small amounts of arsenic and antimony (Vaughan). 
River water may be contaminated by certain discharges from fac- 
tories — as, for example, a river near a chemical manufactory may 
be so poisonous that all fish in its water may die. Chemical poi- 
sons may be purposely' thrown into a well or a stream, arid in this 
manner families or even portions of armies have been affected. 
The method of detecting these poisons will be found in Detection 
of the Common Poisons, page 85. 

Changing Water. — Various gastrointestinal disturbances may 
result from drinking a "new" water, caused, when that water is 



140 LABORATORY METHODS. 

pure, by mineral salts occurring in amount different from that to 
which the alimentary tract is accustomed. 

Goiter Waters. — The authors have had opportunity to investi- 
gate so-called goiter waters in two different localities, and are con- 
vinced that the mineral theories are not well founded. One of 
these villages obtained its water almost directly from the Missis- 
sippi river. No specific amebae were found in any of the many 
samples examined. The goiters occurred only in women who had 
spent most of their lives in the town, and in several of these cases 
were found typical symptoms of Basedow 's disease. In no instance 
was the condition apparent before the age of 12 to 14 years, but 
this "goiter of puberty" usually remained throughout life. All 
women born in the village were not affected, but one who had come 
to the place when 45 years of age developed symptoms at 70. 

Odor of Waters. — The once popular notion that the water with 
no odor was safe for drinking purposes has given place, seemingly, 
to the idea that a foul smell is necessary for a safe and "healthy" 
water. A bad odor may indicate sewage contamination or the 
presence of dead toads, rats, moles, etc., but beyond this has 
little to do with the purity of the water. Other causes of odors 
are: 

1. Pig-pen and grassy odors are due usually to green algae of 
shallow warm waters. 

2. Geranium, oily, and fishy odors are due to certain other algae. 

3. Musty odors are observed sometimes in sewage contamination, 
but are due usually to certain molds. 

4. A hydrogen sulphid odor is due usually to the action of cer- 
tain bacteria on the sulphates. 

5. Putrefactive odors are due not only to dead animals, but to 
decaying vegetable matter in stagnant pools or wells. (See page 
141.) 

Reaction of Water. — This is usually so variable as to be of little 
practical import. 

Detection of Lead. — Symptoms of plumbism may direct suspi- 
cion to the water supply. If lead pipes are used and several peo- 
ple become ill, incrimination of the water is almost inevitable. To 
8 ounces of the sample add iy 2 grains of potassium bichromate. 
A turbidity should result, which in twelve hours will show as a 
slight precipitate on the bottom of the glass vessel. It is best, 
when making this test, to set the vessel on an intensely black back- 



SOME SIMPLE WATER ANALYSES. 141 

ground and observe it at different angles. Control tests with dis- 
tilled water and those containing traces of lead should be made. 

Animal Parasites in Water. — Pinworms and roundworms are 
often distributed by water. The bothriocephalus latus is spread 
by the fish of the Baltic, and has been seen in several American 
clinics. 1 Other forms of tapeworm, trichina, and the several va- 
rieties of the vermes are not usually spread by drinking water, but 
many of the tropical worms are spread in this manner. 

Dead Animals in Water. — The diagnosis of this condition is not 
usually difficult, and the physician is rarely consulted. A drain- 
ing of the reservoir may, however, show no traces of dead animals, 
but the odor may nevertheless persist. Such a water supply need 
not for that reason be condemned, as putrefactive odors arise from 
the anaerobic decomposition of vegetable matter, which points at 
once to stagnation. Connecting the pump nearer the bottom of the 
well or filling up the stagnant space with sand will usually solve 
the problem. (Harrington.) 

Algae in Water. — Vegetable life under water performs the same 
functions as do the higher forms on land, working over animal 
excreta into assimilable matter. If possible, the various algas 
should not be disturbed, as it is an established fact that they will 
destroy sewage bacteria. Some algag give off obnoxious odors, and 
may cause various gastrointestinal symptoms. The cause is not 
usually difficult to find, and has been mentioned in the preceding 
paragraph. Such algEe should not be destroyed by copper solu- 
tions, as is usually recommended, but proper measures should be 
adopted to avoid stagnation. When these small water plants re- 
ceive plenty of oxygen, their presence is more desirable than their 
absence. 

Isolation of the Typhoid Bacillus. — From what has been said it 
seems unwise for the practitioner to attempt any procedure so dif- 
ficult and so discouraging as the isolation of specific pathogenic 
bacteria. The typhoid bacillus has been identified in suspected 
drinking waters by Kubler, Neufeld, Fischer, Flatau, and others. 
In the light of our present knowledge an examination for its source 
is hardly justifiable, save by the expert as a part of a complete 



1 Dr. A. S. Warthin has found the bothriocephalus measle in Lake Superior burbot. 
Indigenous eases of bothriocephalus infection in man have been observed in northern 
Michigan. See Michigan State Board of Health Report, 1912. 



142 LABORATORY METHODS. 

water .analysis. Much work is to be done along this line, and a 
suspected water can be recommended for drinking purposes only 
when the analyst can say, "There are no typhoid germs in this 
water, and under present conditions none can find their way into 
it. " 1 Such a recommendation, coming, however, from an assistant 
in some water laboratory many miles away and from the examina- 
tion of a single sample, is nothing less than criminal. 

Sewage Contamination. — On the other hand, a physician may 
condemn a water as quickly as an expert, provided he give the sub- 
ject a little study and make analyses with sample waters, some of 
which are known to be pure and others to which he has added in 
small amounts sodium chlorid, feces, urine, old manure, etc. 

Sewage contamination may be proven by certain inferences de- 
duced from the sensible combination of two methods. Just which 
of these two has the greater value can not be proven by the counter 
arguments of either chemists or bacteriologists, but by proper con- 
clusions from both. Neither is infallible, and either or both may, 
with certain restrictions, prove contamination by sewage, and the 
latter condition is sufficient to condemn any water. It seems ad- 
visable, in the first place, to eliminate certain examinations usually 
made in water laboratories : 

1. Odor. May be noted, but it is well to remember limitations. 

2. Quantitative determination of certain chemical constituents 
resulting from animal refuse. Where fecal contamination occurs, 
pollution by the urine must be coincident, and urea and sodium 
chlorid will be found. The decomposition of the former leads to 
the formation of various ammonia compounds, the amount of which 
will vary and of which quantitative estimations may be made wdth 
difficulty. 2 Their presence may be overlooked if the estimation of 
the extra chlorin may be easily made and if the value of this esti- 
mation be not excessively limited. (See page 143.) 

3. Certain other considerations of scientific interest, out of no 
practical importance. 

4. Counting oacteria present. This may have some significance 
when the presence of unusually large numbers of microorganisms 



1 The attention of any one interested in isolating the typhoid bacillus from drinking 
water is called to the process devised by Jackson and Melia, where a special agar-agar 
is used in its cultivation. An account of this method was presented to the American 
Public Health Association at Winnipeg in 1908. The process is fully described in 
Leffmann's "Examination of Water," January, 1909. 

2 To quote directly from Dr. Leffmann's preface to his recent (sixth) edition of his 
book on water analysis, we note an important conclusion, "The figures for nitrogen 
or ammonia are of much less value than is generally supposed." 



SOME SIMPLE WATER ANALYSES. 143 

is noted, but even this does not attribute any specific pathogenesis 
to any variety present. 

5. Determination of pathogenic species by animal inoculation. 
This method of bacteriological water analysis, devised by Vaughan, 
is of the highest value when searching for the typhoid and colon 
germs, but can not be considered here. 

Comparative Chlorin — Its Definition. — It is not sufficient to 
draw conclusions from the amount of chlorin in a given water, and 
this is one of the instances where a chemical laboratory at a dis- 
tance fails to give valuable evidence. It is not enough to make a 
quantitative estimation of a certain water, but calculations should be 
made from other carefully selected waters, at least five in number, 
representing all possible sources within a radius of ten miles. A 
careful study of the question will show the necessity for such a pre- 
caution. It would be manifestly unfair to condemn a water in 
certain localities containing much chlorin. The ocean may not be 
far distant, and, moreover, such condition might be explained by 
certain geological formations. An analysis of a water one mile 
distant might give similar results, and it is, therefore, not the 
actual chlorin increase, but the Comparative chlorin content — as 
compared with other waters in that section — of the sample which 
determines its safety. Two wells within a few rods of each other 
may each show large amounts of chlorin, and the question will be 
whether both are contaminated by sewage. Another well a quar- 
ter of a mile away may show considerable less salt, as geological 
formation is not so sharply defined. Samples should, however, be 
taken in other directions before conclusions are attempted, and, if 
the chlorin content of these falls short of those noted in the first 
two wells, it is quite probable that the latter are contaminated; 
if so, compare with results of bacteriological investigations on page 
146. 

Comparative Chlorin — Its Estimation. — At least four waters be- 
sides that suspected should be examined before an opinion is given. 
As the physician may more easily obtain such samples than the 
laboratory at a distance, his opinion will be of greater value. A 
careful selection of these samples may be difficult, but none the less 
imperative. "Level" comparisons as well as "distance" com*- 
parisons are of value, the method of choosing depending on the 
principles outlined above. A study of the geological survey of the 
region in question may be advisable. The results obtained from 



144 



LABORATORY METHODS. 



an investigation of the first set of samples may call for an 
examination of other waters before a final conclusion can be 
reached. 1 The following apparatus and reagents are necessary : 



1. Silver nitrate solution. 2 

2. Indicator^ 

3. Two large tumblers or beakers. 



4. Piece of white paper. 

5. Glass stirring rods. 

6. Buret as used in gastric analysis. 



Place the two beakers side by side on a piece of white paper. 
Add to each exactly 100 cc. of the water and 10 drops of the indi- 
cator. The silver nitrate solution is added drop by drop from the 
buret into the beaker until a slight red tint is noted in the water, 
when it should be compared with the control. This titration is 
conducted exactly as in the gastric analysis, each drop being stirred 
into the water and the reaction given time to take place. After a 
little practice, less than five minutes will answer for the titration 
of each sample. Headings are taken on the buret immediately pre- 
ceding and following the titration. The amount in cubic centime- 
ters of the silver nitrate indicates the number of milligrams of 
chlorin in the water. 

Example. — If 2.5 cc. of silver nitrate are used for the 100 cc. of 
water, this sample contained 2.5 milligrams chlorin (which is a 
very good average in some regions). 

Now titrate in the same manner the other samples, and compare 
results as advised above. Does the sample contain too much chlorin 
— too much chlorin as compared with other waters of that re- 
gion? 

Comparative Chlorin Estimation — Principles Involved. — The 
final red color is caused by the chlorides using up the silver solution 
as long as they are present. A somewhat less affinity is shown by 
the indicator, so that an excess of the reagent causes a formation of 
red silver chromate and proves that the chlorin has been consumed. 

Sources of Error. — The "end reaction," or the first red tint, 
may be difficult for the beginner to recognize. In case there is a 
question, take the reading and then add 1 or 2 drops of the silver 



1 When the worker becomes sufficiently acquainted with the characteristics of the 
drinking waters in his vicinity, and has determined an average chlorin or a standard, 
it will not he necessary in every instance to run control tests. 

- One liter of distilled water contains 4.797 grams of chemically pure silver nitrate, 
and 1 cc. of this solution is equivalent to 1 milligram of chlorin. 

3 One hundred cc. of distilled water contain 5 grams of pure potassium chromate. 
Add some of the silver nitrate solution until a red precipitate forms, and filter. This 
solution serves as indicator. Both solutions must be made up by some reliable firm or 
pharmacist. 



SOME SIMPLE WATER ANALYSES. 145 

solution. In case the end reaction is present, the mixture will be- 
come very red, and it is a good plan to try several "knowns. " 
For example, if 2 milligrams of sodium chlorid are added to dis- 
tilled (chlorin free) water, exactly 2 cc. of the silver solution 
should cause the red tint, etc. 

Value of This Estimation. — From a practical standpoint, 
this computation is of great value. Although a scientific analysis 
would consider ammonia, nitrites, and several other estimations, 
it rarely takes into account the comparison of the different drink- 
ing water sources. As contrasted with the tests to follow (page 
146), chemists and bacteriologists disagree. Neither is infallible, 
and each helps the other. It is a question of personal application, 
and the man who makes both, and then skillfully interprets results, 
gains much mere than he who spends hours in the determination 
of hardness, residue, and total number of bacteria, and who in the 
end can not conscientiously attempt a conclusion. 

Limitations of This Estimation. — The estimation of chlorin 
forms a routine procedure in most water laboratories, and the 
comparative method as described above is not always used. The 
authors have used it successfully in several problems where bacterio- 
logical examinations appeared to be of secondary consideration. So 
far as the simple estimation of chlorin is concerned, and without 
reference to other waters of the region in question, the limitations 
are so many as to render the value and rewards of such labors 
practically nil. 

Bowel Bacilli — Definition. — Some bacteriological method should 
be used in connection with the chemical procedures. Of these 
there are several which, with certain limitations, may be employed 
advantageously by the practitioner. A single principle is involved 
in their application. Where fecal contamination is present, we 
should, without much difficulty, identify certain microorganisms 
whose normal habitat is the bowel. The possibility that these come 
from the alimentary tract of other animals has no bearing on the 
question — a solution of manure was never intended for drinking 
purposes. The colon bacillus, at least, is not a normal inhabitant 
of the gut of a fish (Amyot), and it is the identification of this 
microorganism that indicates contamination by sewage. 

Identification of Colon Bacilli. — Many methods have been de- 
vised, and all have limitations which render them of less value than 
may be desired, yet, when skillfully interpreted, form a valuable 



146 



LABORATORY METHODS. 



adjunct to the comparative chlorin estimation. Two of the many 
methods are presented, with respective cautions concerning inter- 
pretation of results, and either or both may be used: 
Litmus and Gas Method — Apparatus. — 



1. Sterile glucose agar-agar in tube. 
(.See Searching for Germs, page 
38.) 



2. Azolitmin. sterile 1-percent aque- 

ous solution. 1 

3. Platinum loop. 



Method. The tube is inoculated with about 2 cc. of the sample 
water according to the methods described in Searching for Germs, 
page 38. The agar is liquefied and the sterile azolitmin added and 
stirred in, about 5 drops of the latter being usually sufficient. 
When the temperature of the water bath sinks to 120° F., the in- 
oculation must be quickly made. Care should be taken to avoid 
contamination by following the precautions given on page 39. 
With a sterile platinum loop mix well the sample with the rapidly 
solidifying media and keep at body temperature for twenty-four 
hours. Reddish colorations or evidences of gas formation, usually 
both, 'may be caused by the colon bacillus. 

This test is not absolutely conclusive as to the presence of the 
colon bacillus, but is valuable when used in connection with the 
comparative chlorin estimation. The tetanus bacillus may cause 
gas formation, though this hardly occurs within the twenty-four 
hours. The typhoid bacillus may cause the red coloration, but this 
would not argue in favor of the water. 

Fluorescence and Gas Method — Apparatus. — 



1. Sterile glucose agar-agar in tube. 



2. Sterile 1-percent solution of neu- 
tral red. (Griibler.) 



Method. This is a very valuable method, and in a large ma- 
jority of cases, when positive, points conclusively to contamination 
by the colon bacillus. Several anaerobes give the reaction, but the 
typhoid bacillus does not. The technic is identical with that used 
in the litmus test, except that 2 drops of the neutral red solution 
are used instead. The colon bacillus produces in this media not 



1 Certain precautions are necessary when attempting the sterilization of azolitmin 
and other delicate chemicals, as continued boiling results in their partial disintegra- 
tion, and azolitmin may he decolorized, although some of the original color may be 
regained. The colon bacillus should not be present if pure reagents and sterile water 
are used, and neither of these should be contaminated by handling, but a little heating 
is the safest method. It forms no spores, and is usually killed below J. 70° F. — i. e., 
a temperature considerably short of the boiling point. 



SOME SIMPLE WATER ANALYSES. 147 

only gas, but a beautiful lemon-yellow color, with a green fluores^ 
cence. The fact that the bacillus of hog cholera might respond to 
this test does not indicate that such a water should not be con- 
demned. 

Conclusions from a Bacteriological Examination. — Many other 
methods have been proposed both for the identification of the colon 
bacillus and for other microorganisms characteristic of sewage, but, 
used alone, none of them are of much value. When, however, the 
comparative chlorin is high, and when either or both of these 
bacteriological tests are positive, the physician may safely condemn 
the water, and err less frequently than on many other diagnostic 
questions. When the chlorin corresponds with that in other wells 
of that locality (see Limitations, page 145), and bacteriological 
tests are negative, then he may say, "This water is most probably 
safe for drinking purposes." 

Conclusion. — The authors believe that they have proven their 
two propositions, and recommend these examinations to the physi- 
cian. In case assistance is necessary, the expert should not be 
sent a sample of the water, but should be called to the "seat of 
action, ' ' and he should also be called at once in regard to the more 
important sanitary questions previously mentioned (page 139). 

A full and scientific examination of a water usually confuses. A 
sample for a practical analysis should not be sent to a laboratory 
over a hundred miles away for the following reasons : 

1. Certain bacterial changes occur during transportation, even 
where ice is used. (Jordon and Irons.) 

2. An investigation of a single sample by a distant laboratory 
is subject to so many limitations that it is of but little or no prac- 
tical value. 



CHAPTER XIII. 

EVERY-DAY STOOL TESTS. 
Apparatus. — 

5. Glass stirring rod. 



1. Acetic acid, glacial. 

2. Acetic acid, dilute. 

3. Benzidin. 

4. Evaporating dish. 



6. Hydrogen dioxid. 

7. Microscope and accessories. 



Stools are generally so offensive that their examination in private 
practice is often discouraging, and is never necessary as a routine 
procedure. Certain symptoms may, however, direct attention to 
the matter, and it is then that an inspection or a chemical test may 
mean much in a diagnosis. A full and scientific fecal analysis is 
rarely of use outside of the hospital. The physician dreads, and 
with sufficient reason, a necessary test, but an elevated nose never 
diluted any gas. The pathological stool is not usually vile smell- 
ing, while that of the person without ills is almost unbearable. 
Many interesting examinations have been omitted simply because 
they are not, so far as the practitioner is concerned, every-day 
tests. 

Obtaining the Specimen. — A mere inspection cf the entire stool 
is sometimes sufficient — as when looking for large worms, examina- 
tion of infants' stools, etc. — but for chemical and microscopical 
tests small samples containing mucus or bits of blood are selected, 
and these may be carried to the office in a tightly corked bottle, the 
mouth of which is sufficiently wide to permit the entrance of in- 
struments. If a search is to be made for the ameba coli, the speci- 
men must be kept warm until examination of the living animal is 
completed. Bits of material may be transferred to the test tube 
or slide with a' pipette or platinum loop. Such examination may- 
be made after office hours, and suitable methods of deodorizing em- 
ployed. (See page 194.) Smears, after fixation, give off no odor, 



References. — Sahli, Wood, Boston, Webster, Simon, and other works on clinical 
diagnosis. 

148 



EVERY-DAY STOOL TESTS. 149 

and may be examined the following day in case good artificial 
light is not available at the time. 

Odor. — This is of little diagnostic import. There may be little 
or no odor in starvation and certain chronic diseases, but the odor 
of a meat diet is most marked. 

Color. — The following table gives the colors most likely to be met 
in the stool of the adult: 

Color. Etiology. 

Brown, various shades . Normal. 

Dark-brown Meat diet. 

Gray or clay-colored Obstructive jaundice, fatty diarrhea, 

etc. 

Light-yellow Exclusive milk diet, large amounts of 

starches, santonin, rhubarb, senna. 

Red Fresh blood, hematoxylin, in frauds. 

Tarry -black Blood. 

Granular-black Bismuth, iron, and manganese. 

Green Calomel, spinach, lettuce, parsley, in- 
fections with bacillus pyocyaneus, etc. 

Blue or green on standing Methylene blue. 

Violet Santal oil. 

Consistency. — The consistency of an adult's stool is so variable 
as to be of little value in diagnostic procedures, and depends mainly 
on the amount of water present. Fci' the differentiation between 
pus and mucus see The Sputum, page 27. 

Test for Occult Blood. — Dissolve about 1 grain of benzidin in 
about y 2 dram of glacial acetic acid. Stir into y 2 dram of dilute 
acetic acid about 2 grains of the feces, and to 3 drops of this re- 
sulting liquid add 20 drops of 3-percent hydrogen dioxid and 20 
drops of the benzidin solution. Stir the mixture, and, if blood is 
present, a green or blue coloration soon results. This will identify 
very small amounts of blood. Bleeding adenoids must be ruled 
out, and foods containing meat should not be allowed for several 
days preceding the test. It is usually a waste of time to attempt 
the identification of fecal blood by the presence of red corpuscles 
unless they occur in very large amounts. 

Searching for Bacteria. — As a routine procedure, this is of little 
practical value because of the enormous numbers and varieties of 
germs normally present. 1 



1 Of these enormous numbers it seems safe to say that over 75 percent are dead and 
20 percent are attenuated or incapable of growing rapidly in the feces. A few. how- 
ever, notably the colon bacillus, may cause widespread mischief when given a chance. 



150 LABORATORY METHODS. 

Searching for Ameba Coli. — The sample must be kept warm, as 
it is often difficult to identify the amebae, except by the motions of 
their pseudopods, and this activity ceases below 75°-80° F. The 
protrusion of a pseudopod is not a rapid process, and it is not 
usually observed with the eye, but with the aid of a series of rough 
drawings, one of which is taken every thirty seconds. A compari- 
son of these may show that a change in shape has actually oc- 
curred. The amebae may often be differentiated from epithelial 
cells on account of their light-green tint and because they are likely 
to contain leukocytes, bacteria, particles of food, etc., in the cyto- 
plasm. 

Detection of Koch's Bacillus. — The diagnosis of intestinal tuber- 
culosis by the identification of the specific bacillus is subject to 
various limitations, as tubercu-ous sputum may be swallowed, and, 
unless clinical symptoms are sufficient to justify such an examina- 
tion, conclusions may be very misleading. As in the pulmonary 
form, there is a presloughing stage, when few or no bacilli are loos- 
ened into the lumen of the intestine. In acute miliary forms death 
may occur before any ulcers have formed. A positive test for 
occult blood may be the first sign that the mucosa has become 
necrotic, although bacilli usually begin to appear in the feces at 
this time. There are also normally many acid fast bacilli in the 
stools of an adult, notably the timothy bacillus, and these may be 
differentiated from Koch's tacillus by the use of the hydrochloric 
alcohol solution. (See The Urine in Disease, page 123.) The tu- 
bercle bacillus is most likely to be found in the mucous, purulent, 
or blood-stained particles. For its identification see The Sputum, 
page 32. 

Searching for Pinworms. — These may be visible at the anus, 
clinging to the lowest mucosa or hairs, and have the appearance of 
long white crystals of hoarfrost. The first bowel movements are 
most likely to contain them, but never contain their ova. They 
give rise to local disturbances, and may usually be identified with- 
out the use of enemata. 

Searching for Ascaris Lumbricoides. — This occurs in the small 
intestine, but is rarely passed, as it holds fast to the intestinal wall 
by means of spiral turns. "When found, it may be identified by its 
smooth appearance and its light-brown or flesh color. It is usually 
the length of an ordinary lead pencil, and tapers to points at both 
ends. Its ova are often found in the feces, and are elliptical in 



EVERY-DAY STOOL TESTS. 



151 



form, with a hard shell, which is surrounded by an albuminous 
covering'. 

Many physicians desire to save specimens of these worms, and the 
following- technic will give the best results: Kill in 70-percent 
alcohol heated to about 175° F., permit to cool, and transfer to 
preserving fluid made up of 70 parts absolute alcohol, 5 parts 
glycerin, and 25 parts distilled water. 

Searching for Tapeworms. — Segments frequently occur in the 
feces. The ova are globular and are covered with thick capsules. 
It would seem that the morphology is so characteristic that the 






Fig. 33. — Tapeworms compared with vegetable fibers. A, B, magnified segments of 
small tapeworms; C, magnified banana fibers, recovered from stools of an infant. 



confusion of tapeworm segments with other elements would be 
impossible, and it is safe to say that a tapeworm is rarely mistaken 
for anything else. It is, however, often surprising to find how 
closely certain vegetable fibers resemble the segments of small 
teniae, of which the banana furnishes an example. 

Tapeworms may be killed and preserved as follows : Wash in 
water, being careful not to break the worm. Kill in equal parts of 
a saturated aqueous solution of mercuric chlorid and 70-percent 
alcohol, to which has been added 1 percent (of the entire volume) 



152 



LABORATORY METHODS. 



glacial acetic acid. Heat to 160° 1\ and cool. Wash gently in 
running water, and remove excess of mercury with iodin alcohol. 
Rubber gloves should be worn to prevent cysticercus infection. 
Specimens may be preserved as described for ascaris lumbric- 
oides, page 151. 

Tapeworms may be stained and mounted on slides by the follow- 
ing method : After killing, flatten gently between two slides held 
together with a rubber band, and stain in carmine about six hours. 
Remove from between slides and decolorize in alcohol, occasionally 
examining the specimen under low-power objective until a suitable 
picture is obtained. Transfer to 95-percent alcohol and then to 

i — i 1 . . 1 . . . — . 1 — . . 1 . 1 . . . 1 

10 50 100 150 

Microns. 





Fig. 34. — Ova of the most common intestinal -worms compared with a red blood cell. A, 
ovum of hookworm, thin shell containing segmenting or segmented material; B, ovum 
of tapeworm, thick horny shell; C, ovum of round worm, thick shell surrounded by 
albuminous capsule; D, erythrocyte. 



absolute alcohol in order to remove water. Clear in carbol-xylol 
and mount in balsam. If aqueous carmine rather than the alco- 
holic solution is used, it will not be necessary to compress specimen 
until after decolorization. 

Searching for the Hookworm. — The adult parasites are some- 
times found in the stools, but generally a search must be made for 
the ova, which are usually discharged as segmented masses, re- 
sembling a small bunch of large grapes inclosed within a thin shell. 
In cases of "southern anemia," where neither hookworms nor 
their ova are found, a test should be made for occult blood accord- 
ing to previous directions (page 119). This condition is caused 



EVERY-DAY STOOL TESTS. 153 

by the hemorrhage of the mucosa continuing after the hookworm 
has departed to some other portion of the gut. 

Comparison of the Common Ova. — There is hardly any need for 
contrasts when the symptomatology, geographical distribution, and 
other factors are considered. Ova of worms are transparent or of 
a brown color when within the body of the female parasite, but 
when passed by the bowel are yellow or brown because of the uro- 
bilin present. The smallest of these has a diameter three times that 
of the red blood cell, while many are even larger than epithelial 
cells. They should not be confused with the latter because of cer- 
tain characteristics that are absent in all epithelial cells, but some 
of which may be seen in any of the ova. These characteristics are 
their elliptical form of almost geometric regularity, presence of a 
capsule which may in some of the eggs become quite thick, and 
tendencies to segmentation within this shell. 

Fecal Findings in Several Conditions. — 

1. Starvation. Little or no odor ; formed stools. 

2. Vegetable diet. Light-brown color, or even green when fresh 
vegetables are used. 

3. Milk diet. Light-yellow color, with but little odor. 

4. Meat diet. Dark color and very marked odor. 

5. Constipation. The typical stool is very hard, dry, and often 
void of odor ; shows a tendency to crumble. Curiously enough, 
watery stools are sometimes observed in costiveness. 

6. Mucous colitis. Intestinal mucus occurs normally in small 
amounts, but is increased in most pathological conditions. As it 
is easily digested, it comes from the upper bowel only in cases of 
marked diarrhea. Mucus is usually jelly-like in character, but 
may appear as leathery masses, or even as casts of the gut. The 
particles may resemble pieces of asparagus or intestinal worms. 
The mucus of an inflammation is easily differentiated from that 
of a truly nervous condition, as the former contains inclusions cf 
cells or food, while the. latter does not. 

7. Internal injuries. Surgeons watch with interest the first 
urine and feces of emergency cases for signs of blood. 

8. Typhoid. "Pea soup" feces of an offensive odcr, and con- 
taining in small amount pus, necrotic tissue, and sometimes blood. 

9. Tuber cidosis, carcinoma, etc. Blood, pus, etc. 

10. Dysentery. "Watery" feces, containing necrotic tissue, 
amebae, and blood. 



154 LABORATORY METHODS. 

11. Cholera. "Rice water" stools, which late in the disease may 
be practically a suspension of vibrios in blood serum. 

12. Lienteric diarrhea. Much undigested food and little mucus. 

13. Colonic diarrhea. Little undigested food and much mucus. 

14. Drug administration. (See color of feces, page 149.) 

15. Obstructive jaundice. The clay-colored stools are not due so 
much to the absence of bile pigments as to the great increase of 
undigested fats and soaps. 

Stools of an Infant. — Within the first twenty-four hours the 
dark-brown or black meconium should be passed. The stool of a 
milk diet is unformed, of a butter-like consistency, and of a light- 
yellow color. There are many variations from this type, none of 
which can be considered pathological unless symptoms of disease 
or disorder are present, a point that is often overlooked. Any 
variations, however, from the typical milk stool occurring in con- 
nection with illness on the part of the infant become at once of 
the greatest diagnostic and prognostic importance. For example, 
a cheesy stool in a healthy infant means nothing, showing only an 
excess of undigested casein, and does not indicate drugs or diet 
changes, but, should a severe diarrhea accompany or immediately 
follow the appearance of these white, lumpy stools, the indication 
is then clear — barley water or a reduction of proteids. Keeping in 
mind this limitation, the following table is given : 

Appearance of infant's stool. Significance (morbid only when certain 

symptoms justify). 

Pink streak Uric acid infarct (contamination by 

urine). 

White and cheesy Undigested casein. 

Gray Obstructive jaundice, excess of fat. 

Green Changed bile pigments. 

Curds, colic, with constipation or 

diarrhea Proteids at fault. 

Green or greenish-yellow, sour, with 

loose bowels Fats or sugars, usually the former at 

fault. 

Mucus Malnutrition, or accompanies severe in- 
testinal inflammation. 

Red Blood from lower alimentary tract. 

Brown Blood from upper alimentary tract. 

An inspection of the napkin is usually sufficient, and an occult 
blood test may be occasionally indicated. Fatty acids may appear 
as lump-like masses, and are sometimes mistaken for undigested 



EVERY-DAY STOOL TESTS. 155 

casein. The former are, however, quickly dissolved by ether, which 
may be strained away from the stool proper, to reappear when the 
ether evaporates. 

Tampering with the Stool. — The " worm faker," in times past, 
reaped a rich harvest from his victims. When teniae were not ob- 
tainable, earthworms, grubs, and centipedes answered equally well. 
A western ' ' expert ' ' used, with astounding success, a pickled spinal 
cord with its branches, and hundreds of grateful persons passed 
this single specimen. Eventually, however, skeptics began to find 
fault with the darkened bed-chamber, persons became enlightened, 
and vegetable fibers or other bits of extraneous matter were picked 
out and shown under a microscope. The claim has been made that 
certain drugs are capable of dissolving many of these worms within 
the bowel, but often the object is to deceive. 

The pseudo-gallstone seems of late to have become prominent. 
Olive oil or similar fatty bodies are sent to the victim, who is urged 
to take the prescribed substance in large doses, and advised to make 
frequent examinations of the bowel discharges. He soon begins 
to find large soft and greasy lumps, which may be white, but are 
usually green if the bile ducts are not occluded, and consist of fats, 
fatty acids, and certain soaps. The dupe rushes wild-eyed to the 
local physician, who too often agrees that biliary calculi have been 
passed. A second thought, or indeed a brief study of the appear- 
ance of gallstones, would at once expose the imposition. Hema- 
toxylin may color the feces a bright-red, and should cause no more 
concern than a methylene-blue-laden urine. 

Less Frequently Applied Tests. — Certain dietetic tests and dif- 
ficult bacteriological searches, as well as certain scientific examina- 
tions, are omitted, as a study of the various crystals and cells in 
feces may prove a waste of time to the practitioner. 

Value and Limitations of Stool Analysis. — Many of these limi- 
tations have been pointed out, and in selected cases the value of 
certain searches is so well understood that emphasis is unnecessary. 
A routine stool examination in every case of sickness is worse than 
a waste of time to the physician. If, however, certain symptoms 
lead him to suspect the presence of blood or parasites, or the infant 
seems to be slowly starving to death, and he fails to make' the few 
necessary investigations into the character cf the stools, he ceases 
to deserve not only the name "diagnostician," but also "thera- 
peutist." 



CHAPTER XIV. 

TECHNIC OF THE PRIVATE POST-MORTEM. 

Equipment. — A complement of autopsy tools may be selected 
from the surgeon's hand-bag and a carpenter's kit, consisting of a 
cartilage knife or heavy scalpel, amputating knife, large shears, 
probe point shears, surgical saw, bone forceps, chisel, hammer, 
bone drill for Aviring skull cap into place, and wire, sutures, 
needles, probes, tissue forceps, etc. A pair of scales and a foot- 
rule should be included if possible. The high-priced post-mortem 
set offers no advantages over this selection so far as the private 
examination is concerned. 

Private Post-Mortem. — The complete autopsy is rarely con- 
ducted in private practice, and, so far as the various lines of re- 
search are concerned, the clinic will continue to offer greater 
advantages than may be obtained by the general practitioner, but 
this does not indicate that the private post-mortem, incomplete 
though it may be, is wholly without a science. An indiscriminate 
slashing may demonstrate but little, while a few well-selected, 
though crude, dissections may reveal all that is desired to be 
learned. No pathology can be included in this book, except so far 
as elucidation of details in technic is necessary. The description 
given is essentially that offered by Warthin and other experts in 
this branch of medicine, modified only where it is necessary to 
meet the needs of the average practitioner 

Precautions. — 

1. Even though the relatives request the autopsy, a written per- 
mit is always necessary, and such permit may be given by the fol- 
lowing persons in the order named: (1) husband or wife, 
(2) children, (3) mother, (4) father, (5) brothers, sisters, and 
(6) other relatives. 

2. In case of uncertainty as to the cause of death, and if an 
autopsy is forbidden by relatives, the coroner is notified. In case 



References. — Warthin: Practical Pathology; Cattell: Post-Mortem Pathology; Mai- 
lory and Wright: Pathological Technic; Box: Post-Mortem Technic. 

156 



TECHNIC OF PRIVATE POST-MORTEM. 157 

of absence of the latter, the physician, some courts have decided, 
may stop funeral preparations and proceed on his own authority 
to ascertain the cause of death, even though he must resort to an 
autopsy. 

3. No unnecessary mutilation of the cadaver is allowable. If 
necessary, small bits of tissue may be reserved for histological ex- 
amination. 

1. The presence of unprofessional persons is debarred by law, 
but medical students may attend. 

Such, briefly, are the usual legal precautions. In private au- 
topsies, however, certain considerations should be shown the rela- 
tives and friends, who may be waiting for the funeral services to 
proceed, and these considerations may be briefly outlined as fol- 
lows : 

• 1. Provide for the disposal of blood and other waste without 
being seen by the mourners. Suspicious material should be thor- 
oughly mixed with disinfecting solutions and buried. 

2. Perform the work quietly. 

3. Avoid odors by burning coffee, sugar, rags, or tobacco. 

Preparations. — Gowns and rubber gloves should be worn if pos- 
sible, and strict antiseptic methods followed. It is not advisable 
for obvious reasons to use the dining-room table. A work-bench, 
or some rough boards supported by chairs or trestles, answer very 
well. A canvas cot covered with oilcloth has been used, and blocks 
or bricks placed under the legs, but this support is not satisfactory 
when sawing the skull cap. The floor should be covered with 
several thicknesses of paper to avoid soiling. Plenty of cold, clean 
water should be at hand, and warm water is preferable when 
washing up after the autopsy. The cadaver should be entirely 
stripped if possible. For the treatment of wounds obtained during 
autopsies, see Laboratory Prophylaxis, page 179. 

Bacteriological Evidences of Disease. — So much information 
may be obtained by certain examinations during life, that it is 
advisable that these be not deferred until too late to use the in- 
formation in therapeutic procedures, as often nonpathogenic germs 
are already emigrating from the intestines into the tissues before 
the final breath of life leaves the body. 

Microscopic Morbid Anatomy. — Minute inspections are often 
necessary to clear questionable points, and the microscope is then 
called into requisition. Small bits of organs may be dropped into 



158 . LABORATORY METHODS. 

formalin or alcohol, and examined at leisure according to directions 
in Essence of Tissue Diagnosis, page 78. 

Weights and Measures. — Increase of weight accompanies the 
congestion of various organs. Certain atrophies and hypertro- 
phies may cause alterations, often strikingly apparent, but which 
may be evident to the nonexpert only by the proper use of scales 
and foot-rule. To obtain a dimension, thrust a hat pin through 
the desired portion of the organ, and read the result by applying 
the foot-rule. If desirable, measurements may be made with a 
pelvimeter. The following table, taken mainly from "Warthin's 
Protocol," gives average normal weights and dimensions, but the 
English system is used where possible: 

Length of male cadaver 70 inches. 

Length of female cadaver 61 inches. 

Weight of male cadaver 2,344 ounces. 

Weight of female cadaver 1,875 ounces. 

Circumference of skull cap 20 to 30 inches. 

Longitudinal diameter of skull cap 7.5 inches. 

Transverse diameter of skull cap 6 inches. 

Weight of male brain 42.5 ounces. 

Weight of female brain 39 ounces. 

Weight of male cerebrum 36 ounces. 

Weight of female cerebrum 32.5 ounces. 

Weight of male cerebellum 4.5 ounces. 

Weight of female cerebellum 4 ounces. 

Weight of spinal cord About 1 ounce. 

Length of spinal cord About 18 inches. 

Weight of male heart 9.5 ounces. 

Weight of female heart 8 ounces. 

Size of heart. . .Usually that of right clenched fist of individual. 

Size of mitral valve Admits 2 fingers. 

Size of tricuspid valve Admits 3 fingers. 

Size of pulmonary valve Admits 1.5 fingers. 

Size of aorta Admits thumb. 

Thickness of wall of rigbt ventricle 2 to 3 millimeters. 

Thickness of wall of left ventricle 7 to 10 millimeters. 

Weight of left lung 10 to 15 ounces. 

Weight of right lung 11 to 16 ounces. 

Weight of thyroid 1 to 2 ounces. 

Weight of thymus at birth y s ounce. 

Weight of thymus at puberty (disappears after age 25) . % ounce. 

Weight of spleen 5 to 8 ounces. 

Size of spleen 5x2x1 inches. 

Weight of liver 50 to 63 ounces. 



TECHNIC OF PRIVATE POST-MORTEM. 159 

Size of liver 10x12x4 inches. 

Weight of pancreas 3 to 4 ounces. 

Size of pancreas 9x2x2 inches. 

Weight of adrenals 1 to 2 drams. 

Weight of right kidney 4.5 ounces. 

Weight of left kidney Little over 5 ounces. 

Weight of prostate About 5 drams. 

Weight of resting uterus 1 to 1.5 ounces. 

Weight of gravid uterus Very variable. 

Weight of virgin ovaries . 3 drams. 

Weight of adult ovaries 2 drams. 

Cortex of kidney is about % inch in thickness. 

Kidney glomeruli are red and about the size of pin-points, being 
slightly elevated above the surface. . 

Splenic follicles are about the size of a medium brass pin- 
head, gray, and not elevated. 

Uterus wall is about y 2 to 1 inch in thickness. 

Gall bladder wall is about 1 to 2 millimeters in thickness. 

Order of Procedure. — A systematic teehnic is desirable, and, in 
case only portions of the body are to be examined, the following 
order may be accordingly modified : 

1. Inspection, palpation, and percussion. 

2. Head. 

3. Opening of trunk. 

4. Heart is examined before the lungs in order to preserve the 
relation and position of soft clots, etc., which might otherwise be 
drained away. 

5. Lungs. 

6. Neck and thoracic vessels. 

7. Spleen. 

8. Kidneys and adrenals. 

9. Stomach and intestines. 

10. Liver. 

11. Pancreas. 

12. Great vessels. 

13. Pelvic viscera. 

14. Spinal cord. If only the cord is to be examined, it may be 
removed from the back (page 168). 

15. Joints, lymph glands, etc. 

Clinical opinions are of little worth so far as an actual gain in 
medical knowledge is concerned, and the autopsy should be ap- 
proached with the expectation of finding every pathological possi- 



160 



LABORATORY METHODS. 



bility. The circumstances of the case may not, however, permit 
such procedure, as the relatives may consent to an examination 
of only the kidneys for evidences of Bright 's disease, etc. In sus- 
pected Addison's disease or gastric conditions the adrenals and 
stomach should be examined as early as possible, as autolysis often 
rapidly proceeds in these organs after death. 

Inspection, Palpation, and Percussion. — Inspection serves to 
reveal not only the changes due to the cessation of the respiration 
and circulation, but may often show the cause and circumstances 
of death, or give other evidence in regard to the nature of the fatal 
disease. It may be too late for a complete physical examination, 
but a consulting physician may nevertheless be justified in hastily 
palpating and percussing the cadaver. 




Fig. 35. — Scalp incision. The scalp incision must be made in the dorsum in order that 
it may not be visible at the funeral. It extends from ear to ear and completes the 
largest occipital circumference. 



Uncovering the Brain. — The cadaver is laid on its back, and its 
hands placed under the buttocks in order to prevent their dangling 
over the edges of the table. A block should be placed under the 
neck or head in such a manner that the head is raised high from 
the table, the vertex pointing upward. The hair is wetted and 
parted in such a manner as to expose the line of the initial incision. 
The dorsum is selected for the incision in order that no marks can 
be observed at the funeral, extending from ear to ear and com- 



TECHNIC OF PRIVATE POST-MORTEM. 161 

pleting the greatest occipital circumference. If desired, the 
cadaver may be rolled on its belly while the incision is being made 
(Fig. 35), or, if left supine, the head should be raised as directed 
above. 

The cut should extend to the periosteum, and may be made with 
scalpel or cartilage knife. The anterior flap is pushed forward 
and hooked under the chin. The origin of each temporal muscle 
is removed by a semicircular incision and peeled downward. The 
periosteum may be removed prior to opening the skull cap if so 
desired. During these operations and others which may follow 
the prosector should be on the lookout for evidences of disease, and 
hair, scalp, muscles, and periosteum should be subjected to a rigid 
inspection. 

Some operators prefer to remove the entire skull cap by a cir- 
cular cut. The angular cut, however, offers many advantages, 
and is recommended by competent authorities. The angular cap is 
more easily replaced in its natural position, and is less likely to 
present at the funeral a frontal line or joint beneath the skin. 
The angular cut is not nearly so difficult, and is much more cleanly 
because of the absence of hypostatic blood. It may begin at the 
forehead as a circular incision, but is discontinued at the aural 
lines, and from these points two posterior oblique cuts extend 
upward and cross just dorsal to the posterior fontanel, forming 
an acute angle (Fig. 36). 

The left hand may anchor the head of the cadaver with the 
anterior flap, while the right hand uses the saw. A long stroke 
quickly minces the cerebral surface, affording but a short slit into 
the cranial vault, and it is therefore advisable to saw on the most 
convex surface of the bone. A very short stroke is imperative. 

Unless the dura is greatly distended by fluid, or is adherent as 
in babes and the aged, there is no excuse for cutting it. A slight 
loss of resistance indicates the necessity of an exploratory probe. 
The sawdust is white, then red, and again white, as the tables are 
respectively reached, and the dura, when touched, may give forth 
a sound not unlike that of a dry, rustling leaf. "When the probe 
indicates that the sawing has been finished, insert a chisel in the 
right temporal region, making a quick turn. The fingers are in- 
serted and the piece of bone pulled back, the dura stripping slowly 
and always adhering at the longitudinal sinus. 

Examination of Meninges. — To measure the intradural tension. 



162 



LABORATORY METHODS. 



attempt to pick up one fold over the frontal lobe, which picking 
up will be impossible if the tension is increased, and a very loose 
dura indicates atrophy of the brain. The thickness of the dura 
may be measured approximately by inspection; if normal, the out- 
lines of the convolutions and cerebral vessels may be seen through 
it. Examine the superior longitudinal sinus before removing the 
dura. To remove the dura, introduce the amputating knife just 




Fig. 36. 



-Removing the skull cap. The angular method offers many advantages over the 
original circular method. 



to the left of the anterior falx and cut with a sawing motion. Put 
the fingers under the dura and separate oif the vessels ; then cut 
the anterior falx and strip back the dura. 

Removing the Brain.— Place the forefinger of the left hand un- 
der the frontal lobes and pull upward. With a sharp scalpel cut 
each pair of cranial nerves near the exit from the cranial cavity. 
Cut the internal carotid artery and the tentorium with the point 
of the knife sliding along on the crest of the petrous bone, lifting 
up the temporal lobe on either side as this is done. Insert the knife 



TECHNIC OF PRIVATE POST-MORTEM. 163 

through the foramen magnum, severing the spinal cord and verte- 
bral vessels. The brain is laid on the table. 

Examination of the Brain. — The upper surface of the brain is 
examined and then turned over. Carefully examine each vessel in 
the circle of Willis. To open the lateral ventricles, strip away the 
meninges from the top, but leave those on the sides in position in 
order to support the brain substance. With fingers separate the 
hemispheres until the corpus callosum comes into view. Each of 
the lateral ventricles may be opened by inserting the knife without 
injuring the convolutions. To open the third ventricle, cut the 
anterior attachment of the corpus callosum and the lateral pedun- 
cles, and inspect the interior. To open the fourth ventricle, make 
a longitudinal incision through the vermis. To examine the cere- 
bellum, make pig-pen incisions into its substance. To examine 
the cerebral substance, make two sweeping longitudinal incisions 
of cortex and transverse cuts entirely through the pons (pons 
varolii). 

Examination of Special Sense Organs. — Post-mortem examina- 
tions of the eye and ear are rarely made save by experts. After 
removing the brain, the roof of the orbit or of the middle and 
internal ears maj^ be chiseled off and these regions examined from 
above. 

Chief Incision of the Trunk. — This extends from Adam's apple 
(pomum Adami) to the root of the penis (Fig. 37), and is best 
made with a scalpel or cartilage knife. The skin over the neck is 
loose, and must be stretched with the fingers of the left hand as 
the incision proceeds. On reaching the sternum, the belly rather 
than the point of the knife is used, and the cut made deep to the 
bone. At the ensiform cartilage the pressure must be diminished. 
On reaching the epigastrium, a cut is made entirely through to the 
peritoneum and two fingers of the left hand inserted, one on either 
side of the knife, the fingers closely following the path of the knife, 
keeping the tissues taut. The incision is passed around to the left 
of the umbilicus, so that the connections of the latter with the liver 
may not be disturbed. On reaching the pubis the cut is made deep 
to the bone. 

Preliminary Abdominal Inspection. — At this time a brief in- 
spection of the abdominal cavity may be made, noting the appear- 
ance of the peritoneum and omentum, amount and character of 
fluid, position of viscera, distention of gut, and perforations. Take 



164 



LABORATORY METHODS. 



the height of the diaphragm. A transverse abdominal incision 
may be avoided by running the knife under either rectus and cut- 
ting upward to the skin, entirely severing these muscles. If two 
men are at work, one may continue the examination of the ab- 




Fig. 37. — Initial incision. The initial incision of the trunk extends from Adam's apple 
to the root of the penis. 

dominal contents, or otherwise a wet cloth should cover these until 
the thoracic examination has been completed. 

Opening the Chest. — At this time examine the breast by a cut 
underneath, which may be easily hidden. "With well-directed cuts 
peel both flaps of skin and muscle from the ribs as far back as the 
axilla, examining the glands of this region and also the sternum 
and ribs. Beginning at the second rib, cut each costal cartilage 
at its junction with the bony portion of the rib. Cut the dia- 
phragmatic attachments below, and raise the lower end of the 



TECHNIC OF PRIVATE POST-MORTEM. 165 

sternum with the cartilages. With the cartilage knife disarticulate 
the clavicles and first ribs from the sternum. Forcibly separate 
the tissues of the anterior mediastinum and remove the piece. 
Avoid cutting the jugulars, as bleeding would be very profuse. 
Examine the thymus and internal surface of the sternum. 

Examination of the Heart. — This is best examined in position. 




Fig. 38. — Examination of the heart. The heart may he examined in position. A, outlet 
of left pulmonary veins. 

After noting the position of the apex, remove a large portion of 
the anterior pericardium, and you will be ready to open the left 
heart. Seize the apex and swing over to the right, holding tautly 
in this position (Fig. 38), when the left ventricle will become very 
convex. Kun the point of the knife into the apex of the left ventri- 
cle, and cut upward in a line ending at the posterior edge of the 
junction of the left pulmonary vein and left auricle. The actual 



166 LABORATORY METHODS. 

cut, however, stops short of the auriculo-ventricular septum. 
Measure and examine this valve, and then open up the left auricle 
and pulmonary veins. The valvular openings are usually meas- 
ured by inserting the fingers. (See page 158.) The aorta is not 
usually examined at this time. The right heart may then be opened 
by permitting the organ to return to its natural position, and mak- 
ing a cut through the anterior flap, beginning at its edge, and, per- 
pendicular to the cut, extending it toward the right. The cut 
should pass through the greatest transverse convexity, and may 
be continued several inches. Another cut extends upward from 
this to the auriculo-ventricular septum, and, after measuring and 
inspecting this valve, the auricle and pulmonary artery are ex- 
amined. The aorta is measured, and then opened and examined. 

Removal and Examination of the Lungs. — Adhesions are loos- 
ened between the visceral and parietal pleurae if possible, as 
otherwise the parietal layer may be stripped from the ribs. After 
cutting the bronchial and vascular attachments, both lungs are re- 
moved and laid on the table for examination. After a thorough 
inspection of the pleurae, numerous incisions will expose the lung 
substance. Special attention should be given to the apices. Open 
up the larger bronchi and vessels. 

Neck Organs. — These are usually examined in position, and 
rarely offer much in a diagnostic sense. The incision of the thyroid 
will demonstrate the presence of cysts or tumors. 

Examination of the Spleen. — This is seized with both hands, 
and the phrenosplenic ligaments stretched or detached. It is laid 
on the edge of the ribs, and the principal incision made from pole 
to pole. It may thus be examined and returned to its position with- 
out cutting the gastrosplenic omentum. 

Removal and Examination of the Kidneys and Adrenals. — Pull 
over the omentum and intestines to the right. Nick peritoneum 
between descending colon and spinal column, and then lay aside 
the knife, completing w T ith the hands a peritoneal dissection of the 
left kidney and loosening it from its anchorage. Cut the vessels 
on the bodies of the vertebrae, but cut the ureter somewhat lower 
down. The kidney is then removed. Now hold it in the palm of 
the hand in such a manner that the ureter hangs between the fin- 
gers, and cut through the opposite convex surface from pole to 
pole and into the pelvis. Slit the ureters. Attempt to strip the 
capsule, and make other necessary observations. The right kidney 



TECHNIC OF PRIVATE POST-MORTEM. 167 

is removed in a similar manner, and its adrenal is most likely to 
be closely bound to the liver. 

Removal and Examination of the Intestines. — The duodenum 
need not be removed. Separate the stomach and transverse colon 
with the fingers, and then cut transverse mesocolon just back of 
transverse colon, working from below. Tear down the sigmoid 
mesocolon. Pass a couple of ligatures around the gut at the junc- 
tion of the sigmoid and rectum, and sever it between these. The 
hepatic flexure may be torn loose and the ascending colon freed 
from its attachments to the ileum. The small intestine is next 
separated from its mesentery by holding it taut and using the 
knife with a fiddle-bow motion. Pass two ligatures around the 
junction of the duodenum and the jejunum, and divide the gut 
between these. The intestines are now floated in a bucket of cold 
water, opened, washed, and examined. They may be opened with 
shears having one probe point, or with a pair of ordinary shears 
having one point protected with a lead buckshot. 

Examination of the Duodenum. — This is usually examined in 
the body of the cadaver, being easily opened with curved shears, 
carefully avoiding the bile papilla. Squeeze the gall-bladder for 
evidences of stenosis of its duct ; if this is open, the bile should 
enter the duodenum. A finger may be inserted through the pyloric 
orifice to examine for patency. 

Examination of the Stomach. — It is rarely necessary to remove 
this organ in order to examine it. A ligature may be passed 
around the lowest part of the esophagus and the highest part of the 
duodenum, and a longitudinal incision along the anterior wall 
midway between the curvatures will expose all parts of the mu- 
cosa. 

Examination of the Pancreas. — Do not remove it from the cada- 
ver. This is the hardest organ in the body, and a few transverse 
cuts should be sufficient. A piece may be removed for microscop- 
ical study. 

Examination of the Liver. — This may be examined in the body, 
a long incision in the anterior aspect usually sufficing to show all 
macroscopic morbid changes. 

Examination of Retroperitoneal Structures. — Glands, ganglia, 
vessels, etc., may be examined after removing the pancreas and 
mesentery. 

Removal and Examination of the Male Genitalia. — Separate 



168 LABORATORY METHODS. 

the bladder from the anterior abdominal wall with the fingers and 
an occasional use of the knife. Work the hand in behind and under 
the rectum, prostate, and urethra, and cut off the mass, taking 
care to cut the rectum low and the urethra just anterior to the 
prostate. These organs are laid on the table, opened, and exam- 
ined. 

Removal and Examination of the Female Genitalia. — Separate 
the bladder from the pubis as in the male, also dissecting loose the 
rectum from the sacrum with the hand. Separate the cadaver's 
legs and pass two elliptical cuts around the external genitalia, 
following the pelvic outlet, meeting in front at the lowest point of 
the chief incision of the trunk and behind at the back of the anus. 
Dissect upward, keeping the point of the knife close to the bone. 
The mass is removed through the superior strait, laid on the table, 
and the various organs opened and examined. 

Removing the Cord. — In case the examinations of the thoracic 
and abdominal viscera (page 164) are omitted, the cord may 
be removed from the dorsum, as otherwise the ventral method is 
preferable. 

Removing the Cord Ventralward. — In case this is to be done, 
remove all thoracic and abdominal organs, and sponge the cavity 
dry. Beginning at the neck, remove all muscles and soft parts 
from the bone. Place a block under the lumbar region, and with 
a heavy scalpel or cartilage knife cut through the last two inter- 
vertebral disks, using the belly and not the point of the blade, as 
transverse processes protect the cord here. With the bone forceps 
snap the pedicles of the last lumbar vertebra and remove the freed 
body. Cut locse the other pedicles with a hammer and chisel, 
working from below upward, cutting disks and removing bodies of 
the vertebrae as loosened. The spinal roots are put on a tension 
and cut with scissors, when the cord is lifted out. 

Removing the Cord from the Dorsum. — This method is selected 
in case the remainder of the autopsy is omitted, but, on account of 
the hypostasis usually present, it is by no means a clean operation. 
A median incision extends from the hair line to the sacral dimple 
and marks the tips of the spinous processes. The skin flaps are 
then dissected back on both sides. The muscle flaps are made by 
incisions hugging the sides of the spinous processes and dissected 
in a similar manner. These may be reflected outward, and held in 
place by hooks, or may be entirely removed. The lamina? and 



TECHNIC OF PRIVATE POST-MORTEM. 169 

spinous processes are removed with a saw or chisel, or both, and the 
cord is released as described above. 

Autopsy of the New Born. — Several alterations in technic are 
necessary when making the necropsy of an infant. The scalp in- 
cision is identical with that used in the adult, but the bones may 
be separated by cutting their membranous connections with shears. 
Ligatures are passed around the trachea and esophagus before open- 
ing the chest. The lungs and the heart should float if the child 
has breathed. If any lobe of the lung floats, the infant has at least 
gasped. The stomach is opened under water. Bubbles of gas 
usually indicate that air has been swallowed, though they are 
sometimes due to fermentation. 

Completing the Autopsy.— Although notes may be taken, it is 
best to defer the extended discussion of findings. Bones may be 
wired back into place and incisions carefully sutured. Every 
effort should be made to restore the cadaver to its best condition, 
which applies especially to those parts which may be seen at the 
funeral. 



170 



LABORATORY METHODS. 




CHAPTER XV. 



TO FIND THE TREPONEMA PALLIDUM IN SIX MINUTES. 
Apparatus. — 



1. Beaker or evaporating dish. 

2. Flame. 

3. Higgins' carbon suspension. 

4. Medicine dropper. 

5. Microscope and accessories. 



6. Needle. 

7. Physiologic salt solution. 

8. Scalpel. 

9. Slides and cover glasses. 
10. Thermometer. 



Obtaining the Specimen. — Tease lightly the surface of the sus- 
pected mucous patch or cutaneous ulcer, or scrape gently with a 
sharp scalpel the papule or chancre. If the surface of the sus- 
pected syphilelcus seems moist or gummy, this first step may be 
omitted. The fluid, a warm physiologic salt solution, is quickly 
dropped from the capillary pipette on this surface and as quickly 
withdrawn (Fig. 40). An ordinary medicine dropper may be 
drawn out in the flame to serve as a pipette. Four droplets, or 
less, of the solution usually suffice, as the suspension of the trepo- 
nemal will be more concentrated than where larger amounts of the 
liquid are used. As the solution cools very quickly, there should 
be no delay. For practical purposes, a physiologic salt solution 
may be prepared by adding 1 teaspoonful of pure sodium chlorid 
to 1 quart of distilled water, which should be kept warm, but not 
hot. 

Preparing the Specimen. — A clean slide, on which has been 
placed a droplet — not a drop — of Higgins' waterproof black draw- 
ing ink, is ready to receive 1 or 2 droplets of the suspension as 
prepared above. After the ink and droplets are thoroughly mixed 
with a needle or platinum wire, the preparation is ready for ex- 
amination. A cover glass may be added at once and the specimen 



References. — Burri : Wiener Klinische Wochenschrift, July 1, 1909; Williams: 
Archives of Diagnosis, January, 1910, and Journal of American Medical Association, 
December 10, 1910; Barach: Journal of American Medical Association, November 26, 
1910. 

171 



172 



LABORATORY METHODS. 



examined wet. This is not, however, the method preferred in all 
cases by the authors, and, although desiccation often, but not in- 
variably, causes a loss of characteristic windings, a dried prepara- 
tion is more likely to show a typical ideal field. If a dry examina- 




Fig. 40. — Making the suspension. 



tion is desired, no cover glass is to be applied until all evidences 
of moisture are lost. A slight spreading of the droplet is advis- 
able, but this should not be overdone, as otherwise the ideal fields 
will be difficult to find (Fig. 41). When the preparation has dried, 



TO FIND THE TREPONEMA PALLIDUM. 173 

it will show a black center and a small brown peripheral margin. 
The former represents carbon deposits, and the latter consists 
mainly of dried fluid, which contains a very concentrated collection 




Fig. 41. — Spreading the mixture. A, correct spread, lighter areas showing usual loca- 
tions of ideal fields, the hulk of the carhon remaining in the center as the drying 
proceeds; B, droplet spread too thoroughly, with no concentration of the treponemaa. 

of the treponemal That portion of the field last to dry contains 
the most of these germs. 

Examination in the Wet. — By this method the examination may 
be done at once. The treponemae may be observed in motion, and 
the characteristic windings or tnrns of the germs are not lost. The 
examination is done in a strongly transmitted light, and large 
masses of carbon are to be avoided when searching the field. The 
pearly white, almost transparent, treponemae are easily recognized 
on a homogeneous brown or black background, and appear highly 
magnified as a result of this differentiation. 

Examination in the Dry. — When all evidences of moisture have 
disappeared, add a drop of balsam and a clean cover glass. Search 
for an ideal field; i. e., that area which contains the least carbon 
and the most organic matter — not only treponemae, but leukocytes, 
bacilli, etc. This area is usually located at the periphery if the 
spreading has not been overdone. The specific microorganism 
appears much the same as in the wet preparations, except that no 
motions are observed, and the characteristic windings are some- 
times, though not invariably, lost. 

Identification of the Treponema Pallidum. — In order to make a 
positive diagnosis, both wet and dry specimens should be exam- 
ined. The treponema pallidum may be distinguished from spiro- 
chetae, spirilla, cracks, extraneous fibers, etc., by the following gen- 



174 LABORATORY METHODS. 

era! characteristics : a motile corkscrew, much more delicate than 
the other forms ; these turns are close and regular, and appear to 
be somewhat flexible ; its geometric regularity in the spirals, which 
are deeply cut. A thorough study of this microorganism may be 
made either from the literature or from control specimens before 
making a decision. 

The scientific world relies almost entirely on the morphological 
characteristics to identify the treponema, no practical cultural 
methods being known. If any germ or element is found to be 
identical in form and motion with Schaudinn's specific microor- 
ganism, we are then ready to cast aside as useless not only the ink 
methods, but the dark field differentiation method on w T hich the 
expert relies. Nor can the staining time and characteristics be 
termed constants, for in case the treponema pallidum is actually 
found in these inks — and it never has been — the drawing inks may 
then be abolished along with the public drinking cup. (See criti- 
cism of the method below.) 

Sources of Error. — 

1. Use no disinfectant on the syphilelcus for several hours be- 
fore obtaining the specimen. Inquire about the recent use of mer- 
cury or 606, as the authors have noted that either may cause the 
rapid disappearance of the treponemal from the lesion. 

2. Keep the salt solution warm, but not hot. 

3. Use droplets, not drops. 

4. Do not overdo the spreading. 

5. Find an ideal field before attempting to identify the trepo- 
nemal 

6. Study well the characteristics of this germ before reaching 
a decision. 

Disadvantages of the Method. — 

1. Every field is not ideal. 1 

2. Inks are not sterile, and may contain many microorganisms. 
No report has, however, been made that any observer has identi- 
fied the treponema in these inks. 

Criticism of the Method. — The following criticism has been 
made of this method: Certain wavy fibers and cracks appear in 
these inks (Fig. 42), and, while most of them would not receive 



1 To those who prefer to use the India ink as suggested by Burri, the "Chin-Chin 
Liquid Pearl," distributed by Bausch & Lomb Optical Company, is recommended. 



TO FIND THE TREPONEMA PALLIDUM. 175 

very serious consideration by the experienced microscopist, the 
general practitioner is warned against their use. 

Alas, the general practitioner ! Although he has graduated from 
our best medical schools, and usually during the era of microscopy, 
he is hardly given the credit of being able to distinguish a urinary 
cast from a cotton fiber. No critic, however, has yet said, "After 
a careful investigation I can not distinguish fibers and cracks from 
microorganisms." When competent observers can make this state- 
ment, then shall we be prepared to lay aside as worthless not only 
the simple methods, but practically everything that we have thus 
far learned about Schaudinn's specific germ. The treponema has 
not as yet, however, filled all the requirements of Koch's law. 



ABODE F 

Fig. 42. — Treponema pallidum. A, treponema pallidum: B, spirocheta refringens; 0, a 
spirillum; D, cotton fibers; E, flaxen fibers; F, cracks in ink. 

Advantages of the Method. — It is simple, rapid, and the sus- 
pension of the treponemal is twice concentrated, giving in the ideal 
field a maximum number of germs. For the office or bedside 
diagnosis of syphilis the practitioner who has mastered his labora- 
tory courses, and who has kept informed on the treponema pal- 
lidum, may, with a little additional study and practice, depend on 
the method given in this chapter. 

Principle Involved in the Technic. — 

1. There are many treponemal on the surface of every syphilel- 
cus. 

2. These treponemal show a preference for liquids, having a 
tendency to leave their host with the drawing off of certain solu- 
tions properly applied. 



176 LABORATORY METHODS. 

3. Higgins' ink, being a suspension of carbon in a dark fluid, 
permits, when drying, the formation of the ideal nelc. ; which con- 
tains most of the treponemae and but little carbon. 

4. The treponemae are not stained by this ink, but appear as 
pearly, almost transparent, bodies on a homogeneous brown or black 
background, and appear magnified as a result of this differentia- 
tion. 



CHAPTER XVI. 

LABORATORY PROPHYLAXIS. 

The Safe Operator. — The practice of medicine — no less its work- 
shop — is no place for the diffident practitioner. The days of 
"miasms" have long passed, and we know too much concerning 
the subject of bacteriology to hesitate to seek a bit further when 
opportunity offers. We should not, however, despise the "insig- 
nificant little germs." The competent bacteriologist has usually 
passed through at least three stages in his development. In the 
first stage he enters the field trembling with fear, and expects 
disaster from many sources. He is therefore overcautious, and 
consequently nothing goes amiss. After a time, in the second 
stage, he wearies of "details," his fear of microorganisms grows 
less and less, and he often explains his carelessness by "vital re- 
sistance." His evolution to the third stage may be delayed for 
years, but usually takes place when he least expects it. There 
comes a day of awakening, which may end his bacteriological 
career, but which usually ushers in the second reign of caution, 
and he is now qualified to become an expert in this field — in other 
words, he has become a safe operator. 

Germicides, Antiseptics, and Disinfectants. — A germicide is an 
agent which kills germs, an antiseptic prevents or inhibits their 
development, while a disinfectant may do either or both. A brief 
study of the subject will show that distinctions are not sharply 
drawn, but on one point there seems to be a general agreement 
among all authorities — i. e., most chemical disinfectants which 
may safely come into direct contact with the epidermis of the liv- 
ing man are likely to possess very weak germicidal properties. 
Chemical antiseptics are of service only when present, but, when 
removed, those germs remaining are — so far as strictly antiseptic 
action has been realized — freed from any restraining influences. 

Bacteriophobia Versus Carelessness. — The man who is "germ 
afraid ' ' should be debarred from the practice of medicine and from 
the clinical laboratory. On the other hand, the careless diagnos- 

177 



178 LABORATORY METHODS. 

tician is most likely to be a careless therapeutist, and is by no 
means a safe worker for either department. 

Certain precautions were presented in Searching for Germs, page 
42. These admonitions were selected not only from standard 
works, but from actual experiences in the authors' laboratories, 
and are based on the knowledge concerning germs and their habits 
and life processes. The operator may carry in his mind a picture 
of what he is actually doing, and endeavor to explain each step, as 
follows : 

1. He heats to incandescence the platinum loop, thereby destroy- 
ing all germs which may be contaminating it. 

2. He permits it to cool, so that it may not quickly destroy the 
inoculation. 

3. He keeps his hand off it, and does not allow it to touch any- 
thing, so that recontamination may not occur. 

4. He knows that, if it is held in the air, germs may quickly 
alight on it, and he therefore proceeds quickly with his technic. 

5. He touches the point of the wire to the moist colony, and 
knows that some of the many thousands of germs present must 
adhere. 

6. He removes the wire with the captured germs, studiously 
avoiding contamination, and proceeds to inoculate the second tube. 

If he is timid or careless, his technic falls short at some point, 
and his inoculation either fails or, unfortunately, is not made in 
the second tube. 

Special Bacteriological Cautions. — General precautions in re- 
gard to sterilization and safe technic will be found in Searching 
for Germs, page 39. When working in the bacteriological labora- 
tory all wounds on the hands should be protected with bichlorid 
of mercury dressings, or work should be discontinued until an 
aseptic healing occurs. When working with diphtheritic material 
the body orifices, including the eyes, must be studiously avoided 
with the fingers, and the hands should be thoroughly scrubbed and 
rinsed after the examinations are completed. A gargle of an anti- 
septic nature should be used while working with the culture and for 
several days afterward. When much of this work is done, a gargle 
of this character should be kept in the washroom. Special care of 
the eyes should be taken when working with all genital secretions, 
as a single gonococcus in a case of old gleet or leucorrhea may 
cause an acute and destructive ophthalmia. After searching for 



LABORATORY PROPHYLAXIS. 179 

the treponema pallidum the hands should be soaked in a 1: 1,000 
solution of bichlorid of mercury — perhaps the only instance where 
this chemical acts as a germicide. Netting over the window as a 
precaution against the housefly and general cleanliness will make 
the bacteriological workshop many times safer than an indifferent 
use of chemicals — those false securities in mysterious blue solution 
which rarely kill germs. Plenty of light should fall on the labo- 
ratory table. Do not smoke or eat in a bacteriological laboratory. 

Autopsy Prophylaxis. — The routine use of gowns, rubber gloves, 
and general bacteriological measures is, of course, recommended. 
The cold water used during the post-mortem may contain bichlorid 
of mercury, but from this alone safety can not be expected. 

Not only are the acute infections — as diphtheria and typhoid — 
dangerous, but, strangely enough, tuberculosis and actinomycosis 
seem especially prone to attack the prosector. 

Wounds from bone splinters are invariably fatal unless properly 
treated. The vitality of germs powerful enough to survive the 
bactericidal action of the bone would indicate also their great viru- 
lence when coming in contact with living tissues of a weaker class. 
Cases of malignant endocarditis are especially dangerous, and the 
necropsy of such cases should be attended with great care. 

Skin tubercles, or the verrucas necrogenicge, are common, but are 
not to be underestimated, as they have a tendency to spread, 
coalesce, and ulcerate. They should always be cauterized. 

Any wound occurring during a necropsy should be made to 
bleed freely and then cauterized. Bichlorid of mercury dressings 
should be used, and these should be changed at intervals. No 
wound, and especially a deep one, should be covered with adhesive 
plaster or celloidin, as conditions would then become ideal for the 
tetanus bacillus to flourish. 

Burns by Fire. — Burns by boiling water, heated apparatus, or 
even the free flame, are not uncommon in laboratory work. A test 
tube of water which is being heated should always be pointed away 
from the operator, and should be continually shaken to avoid 
' ' shooting. ' ' 

Burns of the first degree should be quickly covered with starch 
paste or soda water, and afterward smeared with petrolatum and 
wrapped with cotton or bandages. If as much as one-quarter of 
the body is affected, the patient should be put to bed immediately, 
and strychnin as well as morphin administered according to indica- 



180 LABORATORY METHODS. 

tions. Such a burn is often followed by a shock or nervous chill, 
and plenty of bed clothes should be at hand. For the treatment 
of shock, reference may be made to standard surgical text books. 
Some fever usually follows a severe burn, for which the treatment 
should be according to accepted practice. 

Burns of the second degree should receive treatment similar to 
burns of the first degree. The blebs may be punctured with a 
hypodermic needle, and the serum aspirated or permitted to ooze. 
The detached epidermis should not, however, be removed at once, 
but only when it shows signs of putrefaction, as it forms a natural 
skin graft and supplies epithelial cells to the new skin. 

Burns of more severe type are treated by cloths wrung out in 
sweet oil, petrogen, or other bland oil. 

Burns by Chemicals. — Mineral acid burns should be washed im- 
mediately with water, which will dilute and remove the acids, which 
may also be neutralized if soapsuds are applied. 

Carbolic acid is not easily removed with water, but alcohol in 
any of its preparations may be quickly applied, removed, and again 
applied, as it dissolves, but dees not neutralize, the poison. After 
a thorough washing with alcohol, a piece of gauze soaked in alco- 
hol may be applied three or four times at intervals of half an hour, 
after which the burn may be treated as if fire, rather than the 
acid, were the destructive agent. 

Burns with alkalies are neutralized with vinegar or other dilute 
mineral acids, and treated as if they were caused by fire. 

Other caustics, as capsicum, may be removed with applications 
of alcohol. 

Explosive Mixtures. — Considerable attention is given to ex- 
plosive mixtures in the discussion of the properties of the various 
reagents (page 186), but it may be well to emphasize here some 
of the more dangerous mixtures of chemicals: 

1. Sulphuric acid with water or anything containing -water in 
considerable quantity. Water should never be added to this acid. 
When its dilution is necessary, add the acid carefully drop by drop 
to the required amount of water. 

2. Potassium chlorate, potassium chromate, and potassium per- 
manganate must not be rubbed up with glycerin, sulphur, tannic 
acid, or other oxidizable substances. 

3. A sudden mixing of strong alcoholic solution with nitric acid 
may cause a quick and severe explosion. 



LABORATORY PROPHYLAXIS. 181 

4. Acids must be added slowly to metals and carbonates. 

5. Hydrogen forms an explosive mixture with phosphorus and 
with air. 

6. Phosphorus should not be permitted to remain in contact 
with the air, but must remain under water. 

7. Sodium and potassium must be submerged in oil, free from 
moisture and the air. 

Inflammable and Explosive Chemicals. — Gases — hydrogen, car- 
bcn monoxid. Volatile liquids — ether, rhigolene, ethyl chlorid, 
celloidin solutions. Nonvolatile liquids — fixed oils and unguents. 
Solids — phosphorus, urotropin. 

Substances Which Should Never be Inhaled. — These include 
the halogens — i. e., chlorin, bromin, etc. 

Arsin must not be inhaled, even in very small quantities, and 
this precaution applies no less to those gases given off when any 
preparation of arsenic is burned. Garlic-like fumes are given off 
by white arsenic when thrown on red-hot coals. 

Hydrocyanic acid is poisonous when inhaled in any quantity, 
even though very dilute, and as the concentration is increased there 
is .likewise an increase in the toxicity, so that the concentrated 
fumes would instantly kill any person exposed. 

The fumes of the strong mineral acids are irritating, and should 
be avoided. 

Hydrogen sulphid and carbcn monoxid gases are poisonous, even 
in moderate quantities. 

If chloroform is used in a room lighted by a free flame, a certain 
amount of chlorin gas is formed, which acts as a corrosive on the 
respiratory passages. 

Sulphuric dioxid and certain of the oxides of nitrogen and car- 
bon dioxid in sufficient quantity are poisonous. 



CHAPTER XVII. 



INDICATIONS FOR LABORATORY AIDS. 



"In which eases shall I resort to the microscope and the test 
tube?" is a question which often arises in the mind of the physi- 
cian. It is not the aim of this work to contrast the value of the 
clinical analysis with other forms of diagnostics, as all these pro- 
cedures are very important branches of scientific medicine and can 
not be properly separated. Either overestimating or underesti- 
mating laboratory aids is the result of ignorance or confusion, but 
it is safe to predict that within the next few years this subject will 
be viewed by the practitioner in a much different light from that 
of the past. 

Below is a list of diseases in which the "mays" and "musts" of 
laboratory aids are indicated, the black type indicating those cases 
where expert assistance is considered necessary. 



Internal psorospermosis — Exami- 
nation of excretions for coceidia. 

Amebic dysentery — Examination of 
stools for amebse. 

Trypanosomiasis — This and all other 
diseases not found in the temperate 
region have been omitted. 

Malaria — Blood smears are treated 
with Wright's . blood stain and a 
search made for the plasmodium. 

Distomiasis — Examination of excreta 
for flukes. 

Tenia — Examination of stools for 
segments or ova. 

Ascaris lumbricoides — Examination 
of stools for adult worms, search- 
ing of muscle bits for encapsulated 
larvae, and inspection of blood 
smears for eosinophilia. 

Ankylostomiasis — Examination of 
stools for the characteristic seg- 



and studies of blood smears for 
evidences of eosinophilia. 

Internal myiasis — Examination of 
excretions or exudations for mag- 
gots. 

Typhoid fever — Ehrlich's diazo re- 
action; blood smears for the study 
of white cells; simplified macro- 
scopic agglutination test with dead 
cultures ; Russo's methylene blue 
test; cultures for typhoid bacilli 
from rose spots, urine, saliva, and 
stools; true macroscopic and mi- 
croscopic Widal tests with living 
typhoid germs. 

Smallpox. 

Vaccinia. 

Varicella. 

Scarlet fever — Repeated urine ex- 
aminations as a prognostic meas- 
ure. 

Measles. 



182 



INDICATIONS FOR LABORATORY AIDS. 



183 



Rubella. 

Mumps. 

Whooping-cough. 

Influenza — Search for specihc mi- 
croorganism. 

Dengue. 

Cerebrospinal fever — Lumbar punc- 
ture and examination of fluid for 
causative germ. 

Lobar pneumonia — A routine exami- 
nation of the sputum is interesting, 
but rarely conducted as a practical 
measure, unless there is reason to 
suspect a tuberculous infection in- 
stead of or in addition to the rav- 
ages of the pneumococcus. 

Diphtheria — Searches for the Klebs- 
Loffler bacillus. 

Erysipelas — Searches for strepto- 
cocci. 

Septicemia — Blood smears may throw 
some light on the diagnosis, espe- 
cially if there is a question in the 
mind of the therapeutist whether 
to use a streptococcus or staphylo- 
coccus serum. 

Sapremia — The germs are usually lo- 
cated at the initial focus, and are 
rarely present in the blood; they 
are saprogenic in character, and 
the culture is usually mixed; it is 
hardly worth while for the prac- 
titioner to attempt to isolate and 
study these, especially as it would 
shed but little light on the thera- 
peutic side of the question. 

Rheumatic fever — Searches for the 
various streptococci; when giving 
the salicylates in heroic doses, occa- 
sional examinations of the urine 
should be made for the presence of 
casts and other evidences of poison- 
ing. 

Yellow fever. 

Hydrophobia — Examination of the 
nervous system of the dog" for the 
Negri bodies. 

Tetanus — A search for the specific 



organism may be. made along with 
therapeutic measures, but the lat- 
ter must never be delayed for an 
absolute diagnosis. 

Actinomycosis — Examination of pus 
for specific microorganisms. 

Syphilis — Searches for the trepo- 
nemal; the various serum reac- 
tions; cytological examination of 
the cerebrospinal fluid in suspected 
syphilitic meningitis. 

Gonorrhea — Searches for the specific 
coccus. 

Tuberculosis — Searches for Koch's 
specific bacillus and for the evi- 
dences of its presence — viz., elastic 
tissue, lymphocytes, etc., according 
to which portion of the body is af- 
fected; section of suspicious lymph 
glands and a study of the patho- 
logical histology; do not attempt to 
demonstrate the presence of the 
specific bacillus in tissues, but try 
to prove the presence of tubercle 
formation. 

Alcoholism, morphine habit, min- 
eral poisons, etc. — Examinations 
of foods, drugs, beverages, excre- 
tions, etc.; isolation of these poi- 
sons in a pure form. 

Food poisoning — Examination cf 
food samples; by "food poisoning'" 
is commonly meant those disturb- 
ances of the A'ital processes arising 
from the ingestion of foods in which 
the poisons or the decomposition 
products of certain bacteria are 
present, which field must be re- 
served for the expert. 

Sunstroke. 

Arthritis deformans. 

Chronic rheumatism — If possible, a 
complete examination of every por- 
tion of the body should be made, 
especially of the tonsils, pleural 
cavities, and other possible sources 
of pus; such analyses may be made 
as are indicated by the findings. 



184 



LABORATORY METHODS. 



Muscular rheumatism. 

Gout — Examinations of the urine. 

Diabetes mellitus — Examination of 
the urine for amount, specific grav- 
ity, general appearance, glucose, 
and diacetic acid; a search of the 
stools for intestinal parasites. 

Diabetes insipidus — Examination of 
the urine for amount and specific 
gravity. 

Rickets — Inquiry into the infant's 
food; if necessary, an analysis of 
the mother's milk or cow's milk, 
and modification accordingly. 

Obesity. 

Stomatitis — Search for the thrush 
fungus, treponema, etc., as may be 
indicated; attempt the detection of 
mercury if such seems to be the 
cause; determination of reaction of 
saliva. 

Diseases of the salivary glands. 

Diseases of the pharynx — Ulcers 
may be examined for pathogenic 
bacteria, especially the tubercle 
bacillus, treponema, fusiform bacil- 
lus, and spirocheta of Vincent's an- 
gina; the diphtheria bacillus and 
pneumococcus may be found in 
healthy mouths, and, unless they 
are present in large numbers, have 
little significance. 

Tonsils — Same as pharynx; the yel- 
low plug of acute tonsilitis is really 
a colony of germs, and may show 
thousands of streptococci or staphy- 
lococci in almost pure culture. 

Diseases of the stomach — Unless 
the diagnosis is evident, it is well 
to supplement it with an examina- 
tion of the stomach contents; add 
to this a test for occult blood 
should the presence of ulcer or can- 
cer be suspected. 

Diseases of the intestine — Proper 
examination of stools, with limita- 
tions in regard to interpreting find- 
ings. 



Appendicitis — A study of the white 
cells, when properly made, gives 
valuable diagnostic and prognostic 
data. 

Intestinal obstruction — Examina- 
tion of vomited material for evi- 
dences of biliary and fecal contam- 
ination; macroscopic inspection is 
often sufficient ; indicanuria is said 
to be present. 

Jaundice — Examination of the urine 
for biliary pigments; inspection of 
stools. 

Diseases of the pancreas — A Cam- 
midge test may be made, but at the 
present time the inferences drawn 
from a positive reaction are sub- 
ject to dispute; tests for urinary 
glucose. 

Peritonitis — In the chronic forms 
where ascites is present an exami- 
nation of the fluid may be made. 

Acute coryza. 

Hay fever. 

Epistaxis. 

Diseases of the larynx — The symp- 
toms and signs of syphilitic and 
tuberculous laryngitis are pathog- 
nomonic, and the collection of ma- 
terial for microscopic examination 
is often attended with difficulty. 

Diseases of the ear — A pus may be 
planted on nutrient agar; a strepto- 
coccus or pneumococcus infection 
is less likely to show a thick and 
creamy pus in large amounts than 
where the staphylococcus is the 
etiological agent. 

Bronchitis, bronchiectasis, asth- 
ma — Sputum examination. 

Diseases of TnE lungs — Sputum ex- 
amination. 

Diseases of the pleura — Examina- 
tion of puncture fluids. 

Diseases of the kidney, bladder, 
etc. — Examination of the urine. 

Diseases of the blood — Partial and 
complete examinations of the blood 



INDICATIONS FOR LABORATORY AIDS. 



185 



for diagnostic and prognostic data; 
in case of pernicious anemia en- 
deavor to discover a cause, for 
which examinations of the urine 
and feces should be made; in case 
of affection of the gastro-intestinal 
tract endeavor to locate the cause 
of chlorosis; in the various forms 
of leukemia, section and examina- 
tion of the lymph glands give but 
little diagnostic data, but where 
leukemia may be ruled out a study 
of these glands may aid to differen- 
tiate tuberculosis from pseudoleu- 
kemia. 

Status lymphaticus. 

Addison's disease. 

Diseases of the spleen. 

Diseases of the thyroid. 

Diseases of the thymus. 

Diseases of the eye — In affections 
of the conjunctiva, associated with 
exudation, smears may be made of 
the serum or pus, and these may 
be fixed, stained, and studied for 
the etiological element — viz. 3 tuber- 
cle bacillus, gonococcus, pneumococ- 
cus, Koch-Weeks bacillus, Morax- 
Axenfeld diplobacillus, streptococ- 
cus, diphtheria bacillus, etc. 

Diseases of the pericardium — Con- 
siderable experience is necessary 
when attempting a puncture ; the 
study of the fluid obtained may 
throw some light on the disease. 



Organic diseases of the heart — ■ 
The sputum may be watched for 
erythrocytes and heart failure cells; 
the urine may be examined for 
casts and albumin. 

Diseases of the arteries — Urin- 
alyses. 

Neoplasms — Section and microscopic 
examination when bits of suspected 
tissue can be obtained; when a hid- 
den tumor is suspected, examine 
the blood for evidences of sarcoma 
(hemoglobinemia) or carcinoma 
( cachectic anemia ) . 

Parasitic skin diseases — Scrapings 
from lesions and examinations for 
parasites; study of the blood smear 
for eosinophilia. 

Diseases of the alimentary tract 
of infant — Analyses of stools or 
vomitus; examination of the ma- 
ternal milk; analysis and proper 
modification of cow's milk. 

Uterine diseases — Study of dis- 
charges; section and examination 
of curettings. 

Pregnancy — Periodical examinations 
of the urine, with special reference 
to the quantity of urea; tests for 
albumose when death of infant is 
suspected. 

Drinking water — Examination for 
evidences of sewage contamination; 
detection of lead from water pipes; 
the more complex problems in 
sanitation. 



CHAPTER XVIII. 

GENERAL INFORMATION. 

For the convenience of the reader, the first portion of this chap- 
ter has been devoted to a study of the physical and chemical 
properties of certain substances. All stains 
and reagents described in this book, as well as 
the more common poisons, are considered and 
alphabetically arranged for ready reference. 



STAINS, REAGENTS, AND OTHEH 
CHEMICALS. 

Acacia — Gum arabic is soluble in 2 parts of 
water, forming a thick liquid; insoluble and 
incompatible with alcohol, ether, oils, mineral 
acids, ammonia, and tartar emetic. 

Acetanilid — Soluble in 2.5 parts of alcohol, 
but almost insoluble in water ; dissolves readily 
in ether and chloroform; incompatible with 
chloral, phenol, resorcin, thymol, and aqueous 
solutions of alkali bromides and iodides. 

Acid, acetic — Possesses the general prop- 
erties of all mineral acids. 

Acid, carbolic — Is not a mineral acid; pos- 
sesses a characteristic odor; is freely soluble 
in glycerin, alcohol, ether, and chloroform; 
soluble in about 20 parts of water and in 2 
parts of sweet oil. 

Acid, chromic — Is really not an acid, but 
chromium trioxid; is soluble in water, but not 
to be mixed with glycerin or other organic sub- 
stances, as it explodes ; incompatible with alco- 
hol, ether, and nearly all mineral salts ; must 
be kept tightly corked and free from moisture. 
186 



Fig. 43. — Five-inch test 
tube (enlarged one- 
third), with various 
amounts of liquid. In 
qualitative work ap- 
proximate amounts 
are always meant un- 
less otherwise em- 
phasized. A, 5 gtt. ; 

B, 1 cc. or 15 gtt.; 

C. 1 3 or 60 gtt. 



GENERAL INFORMATION. 187 

Acid, hydrochloric — A true mineral acid; volatile, and must 
be kept corked ; should be isolated from bottles containing • am- 
monium hydroxid, as the chemical combination of these gases in 
the air results in the formation of solid ammonium chlorid, spread- 
ing a white coating over all laboratory glassware; incompatible 
with alkalies, silver salts, chlorides, chromates, oxides, permanga- 
nates, lead salts, etc. 

Acid, hydrocyanic — An organic compound, soluble in all ordi- 
nary reagents, but used very rarely, as inhalation of its vapor 
causes instant death. 

Acid, nitric — A true mineral acid ; must not be mixed with alco- 
hol. 

Acid, picric — A carbolic acid derivative, soluble in lOOparts of 
water; incompatible with all oxidizable substances, gelatin, albu- 
min, alkaloids, etc. ; explosive with sulphur and phosphorus. 

Acid, sulphanilic — Soluble in warm water, but not in alcohol, 
etc. ; usually purchased ready for the diazo test, but this solution 
may be prepared by dissolving 1 gram of sulphanilic acid in 50 cc. 
of concentrated hydrochloric acid and making up the quantity with 
distilled water to 1,000 cc. 

Acid, sulphuric — A true mineral acid ; water or substances con- 
taining water must not be added to it ; when necessary to dilute it, 
add it slowly to the required amount of water, constantly stirring 
or otherwise mixing. 

Alcohol, ethyl — Freely miscible with water, ether, etc. ; not to 
be mixed with nitric acid, as it will explode; incompatible with 
acacia, albumin, chromic acid, permanganates, etc. ; volatile and 
inflammable. 

Alcohol, fusel (fusel oil) — Mixture of the higher alcohols, es- 
pecially amyl. 

Alcohol, methyl — Volatile and inflammable ; poisonous when 
drank or even when inhaled in considerable quantity. 

Alum-carmine — A nuclear stain, which should be purchased 
ready for use. but may be made up as follows : to 100 cc. of water 
add from 3 to 5 percent of ordinary alum and 1 gram of carmine, 
which is to be boiled fifteen minutes and filtered after cooling, after 
which the solution is again made up to 100 cc. by the addition of 
distilled water to replace that lost by boiling and filtering. 

Ammonium hydrate or ammonla. hydroxid — An aqueous solu- 
tion of ammonia gas ; explodes when suddenly mixed with strong 



188 LABORATORY METHODS. 

mineral acids ; to be kept tightly corked and at some distance from 
the bottle of hydrochloric acid. 

Antipyrin — Easily dissolved in water and the ordinary solvents, 
but incompatible with the majority of reagents and drugs. 

Arsenic trioxid — Soluble in about 5 parts of glycerin; freely 
soluble in hydrochloric acid, strong alkalies, and alkaline carbon- 
ates ; very slightly soluble in water and alcohol ; incompatible with 
salts of iron and magnesium and tannic acid. 

Atropin sulphate — Very soluble in water and alcohol. 

Azolitmin — Purified litmus. 

Benzidin — Slightly soluble in water; freely soluble in alcohol or 
ether. 

Calcium chlorid — Very soluble in water or alcohol; very deli- 
quescent. 

Canada balsam — Should be paper filtered and colorless — not yel- 
low ; is not miscible with water or alcohol, but with the volatile oils, 
chloroform, and ether. 

Carbol-xylol — Add 1 part of pure phenol to 3 parts of pure 
xylol and mix thoroughly. 

Carbol-gentian violet — Should be purchased ready for use in 
ounce bottles. Some workers prefer to use carbol-fuchsin, which 
may also be purchased in liquid form ready for use. 

Celloidin — Shering's celloidin shreds are sold in ounce bottles, 
and the substance is usually kept under water until the solutions 
are needed ; it is freely soluble in alcohol and ether, and the solu- 
tions are highly inflammable; directions for preparing celloidin 
solutions will be found in Essence of Tissue Diagnosis, page 82. 

Charcoal, — Keep well corked, as it readily absorbs gases ; much 
of the charcoal on the market is already thoroughly saturated and 
practically valueless. 

Chloral hydrate — Easily dissolved by all the common solvents, 
but incompatible in alcoholic solutions, and can not be mixed with 
many other drugs ; takes up water easily, and should be kept well 
stoppered. 

Chloroform — Slightly soluble in water, and on account of 
greater specific gravity forms an under layer; soluble in alcohol, 
ether, and volatile oils; must be kept in brown bottles and well 
stoppered, so that its fumes may not come in contact with the free 
flame, as chlorin is thus rapidly formed. 



GENERAL INFORMATION. . 189 

Cocain, hydrochlorid — Freely soluble in water, alcohol, and 
chloroform. 

Copper salts — Soluble in water and alcohol, usually forming 
blue solutions. 

Dimethylamidoazobenzol — Should be purchased ready for use 
in ounce bottles of .5-percent solution. 

Distilled water — Easily obtained at exceedingly low prices 
from druggists ; should be kept well stoppered and in a dry place. 

Eosin, yellow — Should be purchased ready for use in ounce bot- 
tles ; several drops added to a watch glass of distilled water usually 
give a strong stain. 

Ether — Miscible with alcohol, chloroform, and oils; slightly 
soluble in water, the excess forming an upper layer; very volatile 
and inflammable. 

Ethyl chlorid — Usually comes in a tube fitted with a valve for 
releasing; is highly volatile and inflammable; the tube should be 
slightly warmed in order to hasten the flow of the gas, but an ex- 
plosion would quickly follow the application of a high temperature. 

Ferric chlorid — Soluble in water and alcohol; crystals must be 
kept well stoppered and in a dry place. 

Formalin — Formalin is a 40-percent aqueous solution of formal- 
dehyd gas ; miscible in alcohol and water in all proportions ; forms 
explosive mixtures, and incompatible with ammonia, alkalies, tan- 
nin, gelatin, and salts of iron, copper, and silver; to be kept well 
stoppered. 

Glycerin — Miscible in all proportions with water and alcohol; 
forms explosive mixtures with permanganates and other oxidizing 
reagents. 

Gunzburg's reagent — A solution of 2 grams of phloroglucin and 
1 gram of vanillin in 30 grams of alcohol; should be purchased 
ready for use in ounce bottles. 

Haines' solution — A solution of 2 grams of copper sulphate, 
20 grams of glycerin, and 9 grams of potassium hydrate in 175 
grams of water ; test samples by heating — if a reduction takes place 
and a brown precipitate appears, the reagent is to be cast aside as 
worthless ; to be kept in a dark place. 

Hayem's solution — This may be easily prepared by the local 
druggist as follows : mix together about y 2 gram of pure bichlorid 
of mercury, 5 grams of sodium sulphate, and 1 gram of sodium 



190 • LABORATORY METHODS. 

chlorid; dissolve in enough distilled water to make 200 cc, and 
filter if necessary. 

Hemalum — Should be purchased ready for use in ounce bottles. 

Hydrogen dioxid — Miscible in water and alcohol; incompatible 
with most chemicals ; to be kept cool and quiet. 

Iodoform — Soluble in ether, oils, and boiling" water ; incompati- 
ble with balsam of Peru, tannin, salts of silver and mercury, and 
certain other chemicals not commonly used. 

Lead acetate or sugar of lead — Sparingly soluble in water and 
alcohol, but extremely soluble in either when heated to boiling; 
incompatible with many mineral salts. 

Loffler's methylene blue — Should be purchased ready for 
use in ounce bottles; it is a solution of methylene blue to which 
potassium hydrate has been added. 

Marx's fluid — See Vascular Dramas, page 68. 

Mercuric Chlorid or Corrosive Sublimate — Fairly soluble in 
water and alcohol, especially when these have been heated to 
boiling; incompatible with the salts of many metals, tannic 
acid, etc. 

Mercurous chlorid or calomel — Practically insoluble in ordi- 
nary reagents. 

Methylene blue — Soluble in water ; less readily in alcohol ; 
should be purchased ready for use in liquid form in ounce bottles. 

Methyl green — Aqueous solution containing 1 percent acetic 
acid. 

Morphin sulphate — Soluble in water, especially when hot ; 
sparingly soluble in alcohol; incompatible with salts of metals, 
alkalies, iodides, tannic acid, etc. 

Nessler's reagent — A solution of 10 grams of potassium iodid, 
5 grams of mercuric chlorid, and 32 grams of potassium hydrate in 
distilled water to make 200 cc. 

Neutral red — Soluble in alcohol or water. 

Oil spearmint — A true volatile oil. 

Opium — Soluble in alcohol, which, when in excess, permits the 
addition of a certain amount of water without precipitation; in- 
compatible with alkalies, silver nitrate, and alkaloidal precipitants. 

Phenacetin — Fairly soluble in alcohol, but hardly in water. 

Phenolphthalein — Easily soluble in alcohol, but not in water 
unless small amounts of alkali are added. 

Phosphorus — Slightly 'soluble in alcohol, especially when the 



GENERAL INFORMATION. 191 

latter is hot, but almost insoluble in water ; soluble in the fixed oils 
and freely soluble in carbon disulphid. 

Potassium chromate — Soluble in water, but not in alcohol. 

Potassium bichromate or potassium dichromate — Soluble in 
water, especially when the- latter is heated; practically insoluble in 
alcohol. 

Potassium hydrate — A caustic alkali easily dissolved by water 
or alcohol. 

Silver nitrate — Very soluble in water and fairly soluble in alco- 
hol ; soluble in 5 parts of distilled water ; incompatible with salts of 
metals, tannic acid series, organic substances, and aqueous solutions 
of vegetable drugs ; keep in brown bottle protected from light. 

Sodium chlorid — Very soluble in water, but almost insoluble in 
alcohol; incompatible with alcohol, silver nitrate, and certain salts 
of lead and mercury. 

Sodium citrate — Exceedingly soluble in water; slightly soluble 
in alcohol. 

Sodium nitrite — Easily soluble in water ; slightly soluble in 
alcohol ; incompatible with many chemicals ; to be kept well stop- 
pered. 

Strychnin sulphate — Fairly soluble in water, especially when 
the latter is heated or glycerin is added. 

Stannous chlorid — Very soluble in water ; fairly soluble in alco- 
hol ; to be kept well stoppered. 

Sulphanol — Fairly soluble in hot Avater or alcohol, but spar- 
ingly when these solvents are very cold. 

Sweet oil — A true fixed oil, which becomes rancid on exposure 
to light and air. 

Thionin — Soluble in water, but concentrated solutions usually 
require filtration at intervals. 

Tincture turmeric — An alcoholic solution of curcuma ; to be 
kept in a dark place. 

Urotropin or hexamethylenamine — Soluble in water, but not 
in other reagents ; tablets burn in the air with a hot, colorless 
flame. 

Zinc — This must be chemically pure, or at least arsenic free; 
should be purchased in the form of short sticks; zinc dust to be 
used when making up Meyer's blood test solution; to be kept well 
stoppered. See The Urine in Disease, footnote on page 119. 

Wright's blood stain — Should be purchased ready for use in 



192 



LABORATORY METHODS. 



liquid form in ounce bottles; to be discarded as soon as a precipi- 
tate forms ; precipitation to be avoided by controlling evaporation ; 
to be kept well stoppered. 



WEIGHTS AND MEASURES. 

Apothecaries' Weight. 

20 grains = 1 scruple. 

3 scruples == 1 dram. 

8 drams = 1 ounce. 

12 ounces = 1 pound. 

Apothecaries' Measure. 

60 minims :— 1 fluidram. 

8 fluidrams = 1 fluidounce. 

16 fluidounces = 1 pint. 

8 pints = 1 gallon. 

Equivalents. 

1 grain = 60 milligrams. 

15 minims = 1 cubic centimeter. 

1 5 grains .,. = 1 gram. 

1 dram — 4 grams. 

1 fluidram = 4 cubic centimeters. 

1 fluidounce = 32 cubic centimeters. 

1 ounce — 32 grams. 

1 quart — 1 liter. 

A teaspoon contains about 1% fluidrams, a dessert spoon about 
2y 2 fluidrams, and a tablespoon about 5 fluidrams. 



MISCELLANEOUS. 

Drop Method for Preparing Approximate Percentage Solutions. 

— In qualitative analysis (but not in therapeutics) we may, when 
considering aqueous solutions, use the terms "drop" and "minim" 
synonymously, and this method gives fairly accurate percentage 
solutions. For example, suppose a 10-percent solution of silver 
nitrate is desired. This substance is soluble in one-half its volume 
of water, or each drop of a concentrated solution in distilled water 
contains approximately 2 grains; or, to state it in other terms, to 
each of these drops must be added 19 drops more of the water to 



GENERAL INFORMATION. 



193 



obtain the desired percentage. This method does not, however, 
answer for quantitative tests, nor for mixtures intended for thera- 
peutic purposes. 

Fahrenheit and Centigrade Equivalents. — On the Fahrenheit 
thermometer the space between freezing and boiling points is di- 
vided into 180 equal parts or degrees, the zero point being 32 de- 
crees below the freezing of water. On the Centigrade thermometer 
the space between freezing and boiling points is divided into 100 
parts or degrees, the zero point being at the freezing of water. A 
degree of Fahrenheit is therefore % degree of Centigrade. The 
following are simple rules for the conversion of temperature of 
Fahrenheit to Centigrade and Centigrade to Fahrenheit : 



ABOVE 32° F. AXD 0° C. 
Fahrenheit to Centigrade. — Subtract 32, multiply by 5, divide 



by 9. 



Centigrade to Fahrenheit. — Multiply by 9, divide by 5, add 32. 



Fahrenheit to Centigrade. 
50° F. 
32 


Centigrade to Fahrenheit 
10° C. 
9 


18 
5 


5)90(18 
32 


9)90(10° C. 


50° F. 



The examples show that 50° F. above 0° equals 10 C.° above 0°, 
and vice versa. 

BELOW 32° F. AXD 0° C. 

Fahrenheit to Centigrade. — Subtract from 32, multiply by 5, 
divide by 9. 

Centigrade to Fahrenheit. — Multiply by 9, divide by 5, sub- 
tract from 32. 



Fahrenheit to Centigrade. 
32 
5° F. 



9)135(15° C. 



Centigrade to Fahrenheit. 
15° C. 
9 



5)135(27 from 32 
27 



5° F. 



The examples show that 5° F. above 0° equal 15° C. below 0°, 
and vice versa. 



194 LABORATORY METHODS. 

Spoon Urinalyses. — These analyses have been popular with the 
general practitioner as bedside tests, and give fairly good results, 
but are not to be compared with the urinalyses described in this 
book (page 111). The following tests are quoted from Richter, 
as the necessary apparatus may not be at hand, but they would not 
be accepted by any insurance company nor by any high authority 
in a consultation. 

To test for albumin, take a half teaspoonful of urine, add a small 
pinch of salt, and heat over a lamp or match. When it begins to 
steam and bubble, add a few drops of vinegar and watch for pre- 
cipitate. 

To test for glucose, dilute 1 or 2 drops of the urine with a few 
drops of water in a spoon. Carefully evaporate to dryness with a 
little heat. Now again slowly heat, when almost suddenly a char- 
acteristic orange-brown spot and an unmistakable odor of caramel 
will prove the presence of sugar. One-third of 1 percent is easily 
detected in this manner. Urine free from sugar colors a smoky 
black, and on further heating emits its peculiar urinous odor. 

Deodorizers. — Previous references have been made to this sub- 
ject and certain formulas presented (page 24). The burning of 
sugar or rags, as practiced by the housewife, is efficient, not only 
because of its "counter odor," but for the reason that the carbon 
thus formed in its heated or nascent state readily absorbs obnoxious 
gases. Foul odors from certain substances may be covered up by 
certain volatile oils, notably cassia and bergamot. 

To Remove Rust from Instruments. — Broder's method will be 
of value to the surgeon as well as the analyst who desires that his 
outfit present the best possible appearance. Fill a suitable vessel 
with a saturated solution of chlorid of tin in distilled water, im- 
merse the rusty instruments, and let them remain over night. Rub 
dry with chamois after rinsing in running water. 

To Remove Stains from Fingers. — Anil in dye stains may be 
quickly removed from the fingers by washing with a cloth damp- 
ened in a solution of 4 parts alcohol and 1 part ether. 

Mineral acid stains and those caused by potassium permanganate 
may require several good scrubbings in hot soapsuds before they 
disappear. 



APPENDIX. 

Being a concise report of recent and more special laboratory methods which 
by virtue of their simplicity and usefulness will appeal as practical to the 
man who desires to go somewhat more deeply into this enticing department 
of medicine. 

Bedside Estimation of Urinary Acidity. — Reference has been 
made to the significance and estimation of urinary acidity (pages 
109 and 123). To avoid the tedious buret method of titration, 
Harrower has devised a simple instrument in the form of a gradu- 
ated test tube of heavy glass. This may be carried in the phy- 
sician's handbag and the estimations made at the bedside of the 
patient. The physician has only to fill the tube with the urine 
up to the 10 cc. mark, add a couple drops of phenolphthalein so- 
lution and then the decinormal sodium hydrate solution drop 
by drop, shaking well after each addition until the first perma- 
nent pink is obtained. The percentage is then read off directly 
from the scale on the side of the tube. By keeping track of the 
amount of urine passed daily, estimation may be quickly made 
of the total quantity of acid excreted in that time. 

Significance of Indicanuria. — Indican is not a normal constitu- 
ent of the urine but occurs as a result of proteid decomposition 
at some point in the body. Thus in the purulent conditions, e. g., 
hidden abscess, the presence of indican in the urine may lead us 
to diagnosticate the condition. In malignant growths where a 
considerable degree of degeneration and necrosis is present, in- 
dican may appear in the urine in large amounts ; but if we can 
rule out these two sources of indican, we have still to reckon with 
another of considerable importance ; viz., bacterial decomposition 
of proteid foodstuffs in the colon. In other words, by the de- 
tection of indican in the urine of a person in whom hidden pus 
or other degenerative changes are unlikely, we gain easily and 
quickly evidences that affairs in the large bowel are not as they 
should be, and that it is time that this natural incubator be 
cleaned out. 

195 



196 APPENDIX. 

The finding of excessive amounts of acid in the urine furnishes 
further proof, but if such is not present, we are scarcely justi- 
fied in concluding at once that the indican does not come from 
the colon. The acids accompanying such a condition may be 
well cared for by the neutralizing precursors of urea; but noth- 
ing is present to render inactive the telltale indican. 

While it is not the aim of the authors to delve deeply into the 
mysteries of indican chemistry, another word must be added to 
complete the diagnostic and prognostic consideration of this 
body. INDICAN IS NOT A NORMAL CONSTITUENT OF THE 
URINE, let it be repeated. In case pyogenic processes in the 
tissues (pyorrhea alveolaris, otitis media, septic tonsils, sinusitis, 
tuberculous cavities in the lungs, empyema, gall bladder abscess, 
chronic appendicitis, salpingitis, pyelitis and chronic gonorrheal 
prostatitis) neoplasms and true copremia are finally ruled out, 
we have not yet finished our search for the source of urinary 
indican. Few of us are entirely normal: a gastric hypoacidity 
may interfere with the proper digestion of the proteids ; or we 
may be taking too much proteid food. In such cases, however, 
the indican usually occurs in merest traces, so that a consider- 
able amount of this substance always means a strictly pathological 
condition. For copremia is not to be regarded lightly, it being 
quite possible that the poisons absorbed from the bowel are 
highly toxic to the kidney tissue and may have much to do with 
the etiology of Bright 's disease. 

Furthermore, a mere cleaning out of the colon may not cause 
a disappearance of the urinary indican; but such circumstance 
does not prove that the condition is not copremia. The same 
microorganisms quickly proliferate to numbers equal to those 
first attracting our attention. The bowel must be kept reno- 
vated and a new flora (e. g. — the Bulgarian bacillus) introduced. 
Reduce also the proteid diet, and if these measures fail to re- 
duce the indican, it is very likely that proteid decomposition in 
the tissues is responsible for the indican. This substance occurs 
in large amounts in enteroptosis inasmuch as the bowel bacilli 
have ample opportunity to proliferate and decompose the pro- 
teids. The same may be said concerning intestinal obstruction 
or fecal-clogged colon. 

Detection of Indican. — The following method has been recom- 
mended by Dr. Harrower and others, and is simple as well as 



APPENDIX. 197 

reliable: To one dram of the urine, add one dram of pure hy- 
drochloric acid, one half dram of chloroform and two or three 
drops of reliable hydrogen peroxide. Shake well for a minute 
and set aside. If a reaction fails to appear, shake again and 
set aside. The quantities of liquids used need not be accurate, 
but should be approximately so. 

The chloroform dissolves the indigo which has been liberated 
by the action of the acid and peroxide ; and as it sinks to the 
bottom of the tube, a blue or violet color is noted, the intensity 
of the tint varying directly with the amount of indican in the 
urine. Thus we speak of a trace, a small amount, a moderate 
amount, a large amount, and a tremendous amount — although 
we have no very reliable method of estimating the degree of in- 
dicanuria. Sometimes a permanent red color may take the place 
of the blue in the chloroform, and this is called red indican. Red 
indican probably always comes from the bowel, bearing a relation 
to skatol not unlike that which indican bears to indol. It is 
more likely to arise from the decomposition of vegetable protcids 
in the colon; and such patients are usually irritable whereas 
those with blue indican are depressed. 

Sources of Error .—Inquire concerning the administration of 
potassium iodid, salicylic acid and urotropin as these may give 
pseudoreactions. Such drugs should be omitted before testing 
the urine. Iodid of potash imparts to the chloroform a rose-red 
color ; but this may be driven away by shaking with a few drops 
of alcohol or sodium hyposulphite, whereas the true red indican 
is refractory to these reagents. 

The urine should be fairly fresh and should contain no pre- 
servatives. Often the supernatent liquid becomes of a dark 
color, but the chloroform does not seem to be able to dissolve this 
indigo. In such case, add one-half dram of alcohol when upon 
shaking, solution will be effected by the chloroform-alcohol 
mixture and the heavy layer comes down blue or violet. 

Detection and Significance of Indolacetic Acid. — Many of the 
poisonous acids formed in the colon and excreted by the kidney, 
baffle accurate identification by tests applicable in the physician's 
laboratory. A notable exception is indolacetic acid which may or 
may not be formed in the copremic process. Its significance is 
the same as that of indican, though either may be present with- 
out traces of the other being found. To test for this substance, 



198 APPENDIX. 

add to about one dram of the urine, one drop of a one percent 
solution of potassium nitrite and a few drops of hydrochloric 
acid. If positive, a pink color is noted, which varies in intensity 
directly with the amount of indolacetic acid present. As soon 
as the color is or is not noted, the test may be finished for indican 
with the same sample as directed above. 

Significance and Detection of the Bence-Jones Body. — The 
finding of this substance in the urine may be regarded as 
pathognomonic of multiple myelomata. Exceptions have been 
claimed, but these must be very few in number. Acidify the 
urine if necessary. If it is not clear, filter it. Heat a specimen 
in a test tube. When it becomes quite warm — almost hot — a 
marked turbidity or cloudiness may be observed which grows 
more and more intense as the temperature approaches boiling; 
but as ebullition begins, the urine begins to clear; and when it 
boils the cloudiness wholly or nearly disappears. Now if the 
contents be allowed to cool, the turbidity reappears. 

Sulphosalicylic Acid Test for Urinary Albumin. — Physicians 
are loath to carry nitric acid in their handbags ; and in many 
cases a strong aqueous solution of sulphosalicylic acid may an- 
swer equally well. This reagent should not be concentrated, in- 
asmuch as many crystals are deposited in the bottom of the vial 
and may interfere with the test. It is well to keep the vial 
tightly stoppered as evaporation appears to encourage the depo- 
sition of these crystals. 

This is a very sensitive test for serum albumin. It is to be 
carried out at room temperature — i. e., the urine and the reagent 
are not to be heated. To one dram of the urine (filtered if not 
clear) , add a couple of drops of the reagent and shake sufficiently 
to mix. Hold in the light and watch for the appearance of a 
cloudiness. This usually manifests itself promptly, but may be 
delayed. In case of question, add a few more drops of the re- 
agent. Rarely more than five drops will be required for a dram 
of urine. Compare with some of the urine to which the reagent 
has not been added. 

This test may be applied to a clear, unfiltered urine or to one 
which comes through the filter clear; and this statement applies 
to most urines, although occasionally a specimen cannot be 
cleared by filtration. In such contingency, the test is scarcely 
applicable and we must fall back upon the older methods. 



APPENDIX. 199 

Hermann-Perutz Serum Test for Syphilis. — At present the 
Wassermann. is denied the busy practitioner, as considerable time 
and experience is necessary for its completion. The Hermann- 
Perutz reaction was devised to fill this want ; and though a new 
test, it bids fair to find a permanent post in the physician's labo- 
ratory. This reaction requires for its completion but three ele- 
ments, viz. — 

1. Reagent A. 

Sodium glycocholaie 2.0 

Cliolesterin 0.4 

Ninety-five percent, alcohol 100.0 

. 2. Reagent B. 

A two-percent aqueous solution of sodium glycocholate. 
3. Specimen for examination. 

Fresh blood serum from the patient. 

The reagents should be prepared just before the test is set up. 
Some blood from the patient's ear or finger is collected in a 
sterile test tube and slanted in an icebox over night. (A vacuum 
bottle with some bits of ice will serve equally well.) In the 
morning the serum is collected by means of a sterile pipet and 
placed in a sterile test tube; and a sterile cotton plug is placed 
in its mouth. The first tube and clot are of no further use in the 
test. The serum is now "inactivated" by placing in an incubator 
(or vacuum bottle) at 132-133 degrees F. for thirty minutes. 
Before the test is set up, dilute Reagent A with sterile distilled 
water in proportion of 1 :20. 

In a third small, sterile test tube mix .4c. c. of the serum and 
.2 cc. of each reagent. These quantities may easily be taken up 
by the aid of an accurately graduated pipet. (The authors have 
obtained excellent results with 5 drops of the serum and 3 drops 
each of the respective reagents; but the original method offers 
no great difficulties if the physician secure a proper pipet.) The 
mixture is vigorously shaken and then set aside at room temper- 
ature, providing the room does not become chilled or overheated. 
The tube is examined -at frequent intervals. If the reaction is 
positive, a flocculent precipitate will appear. A control should 
always be carried out alongside this test, with a serum known to 
be nonsyphilitic. 

Value and Interpretations of the Hermann-Perutz Reaction.^ 



200 APPENDIX. 

We believe that the meaning of the positive reaction is identical 
with that of the Wassermann. If the reaction is positive, the 
patient has syphilis: if it is negative, we are not certain as to 
the diagnosis and there is still a possibility that he has syphilis. 
The disappearance of the reaction under mercury or arsenic is 
not proof sufficient that the p'atient is finally cured, although it 
probably shows some degree of success in this direction. Thus 
only the positive reaction is of value and this value is diagnostic 
only; and so far as we have conclusiye proof is never prognostic. 
Apparently the reaction is as sensitive as the Wassermann; and 
is doubtless as dependable. Because of its comparative sim- 
plicity, it bids fair for a place among our valuable laboratory 
methods, not only so far as the needs of the general practitioner 
are concerned, but by virtue of this very simplicity, the sources 
of error are reduced to a minimum. Carried out in connection 
with control serums and with the butyric acid test (see page 
99) its diagnostic worth is great indeed. 

Recent Studies in the Painful Oxalurias. — Several months ago 
one of the authors in a review of the oxalurias, 1 showed that cer- 
tain cases of lumbar distress and hematuria may be explained by 
the mechanical injury to the mucous membrane of the urinary 
passages, occasioned by the passage of sharp crystals of calcium 
oxalate ; the symptoms increasing in intensity as larger numbers 
of these crystals are passed. Such a condition may be termed 
painful oxaluria, oxaluria dolorosa, and is by no means an infre- 
quent one, though often missed by the practitioner. The treat- 
ment is dietetic and medicinal, never surgical unless other trou- 
bles complicate. The practitioner should remember that the crystals 
of calcium oxalate are very small; and this applies especially to 
those causing symptoms. They cannot be identified by the low 
power objectives; and are often missed by the higher powers if 
the worker is careless. There are other forms of oxaluria, and 
these are not associated with symptoms; but here the crystals 
are neither deposited in such large numbers nor so high in the 
urinary tract; they are smaller and their edges not so sharp and 
their excretion does not extend over long periods of time. Inas- 
much as these crystals are often found in fermenting urines, a 
fresh sample should be examined before an opinion is ventured. 

1 Medical Record, May 11. 1912. 



INDEX, 



A 

Abortifacients, composition of, 87 
Acacia as a reagent, 187 

for tissue sectioning, 80 
Accessories for laboratory, 14 
Acetanilid, detection of, 88 

properties of, 186 
Acid, acetic, 62, 186 

carbolic, 186 

chromic, 186 

hydrochloric, 75, 181 

combined, estimation of, 75 
free, tests for, 75 

hydrocyanic, 187 
cautions, 181 
detection of, 91 

lactic, 75 

nitric, 110, 187 

picric, 187 

sulphanilic, 100, 187 

sulphosalicylic, 198 

sulphuric, 135, 187 
Acidemia, 123 
Acidosis, 123 
Acids, detection of, 88 
Actinomycosis, 49 

laboratory aids in, 183 
Addison's disease, 185 
Albumin in urine, 109 

in sputum, 29 
Alcohol, 187 

amyl, 187 

detection of, 88 

ethyl, 187 

methyl, 187 

fusel, 187 
Alcoholism, laboratory aids in, 183 
Algae in water, 141 
Alkalies, detection of, 88 
Ameba coli. 150 
Amebic dysentery, 182 
Ammonia. 187 

detection of, 88 
Anemia, 58 
Ankylostomiasis, laboratory aids in, 

182 
Antiseptics, definition, 177 
Apothecaries' weight, table of, 192 

measure, table of, 192 
Apparatus. 24 

cabinet for, 24 



2U1 



Apparatus — cont'd. 
for autopsies, 156 
for bacteriological tests, 37 
for blood analysis, 53, 57 
for diazo test, 101 
for gastric analysis, 71 
for Marsh test, 85 
for milk analysis, 125 
for poison tests, 85 
for sputum analysis, 27 
for stool tests, 148 
for treponema test, 171 
for urinalysis, 107 
for water analysis, 144, 146 
for Widal reaction, 104 
Arsenic, 188 

detection of, 89 
Arthritis deformans, 183 
Ascaris lumbricoides, 150 
Ascites, 97 
Atropin, 188 

detection of, 90 
Autopsy prophylaxis, 179' 
technic, 156 

bacteriological, 157 

brain, 162 

chest, 164 

cord, 168 

duodenum, 167 

equipment for, 156 

female genitalia, 168 

heart, 165 

in new born, 169 

inspection, palpation, and per- 
cussion, 160 

intestine, 167 

kidneys, 166 

liver, 167 

lungs, 166 

male genitalia, 167 

meninges, 161 

microscopic morbid anatomy, 157 

order of procedure, 159 

pancreas, 167 

precautions in, 156 

preparations for, 157 

private autopsy, 156 

prophylaxis in, 179 

special sense organs, 163 

spleen. 166 

stomach, 167 



202 



INDEX. 



Autopsy technic — cont'd. 
trunk, 163 

uncovering the brain, 160 
weights and measures, 158 

Azolitmin, 188 

in water analysis, 146 

Azoospermatism, 97 

B 

Bacteria, 37 

in blood, 67 

in exudates, 97 

in feces, 149 

in stomach contents, 75 

in urine, 121 

in water, 142, 145 
Bacteriophobia, 177 
Bad whisky, composition of, 87 
Bass Test, 104 
Bence-Jones body, 198 
Benzidin in blood tests, 148 

properties of, 188 
Betaoxybutyric acid, 116 
Bilirubin in urine, 113 
Bitters, composition of, 87 
Blenorrhea, vaginal, 98 
Blood, 53 

apparatus for examination of, 53, 57. 

artifacts in, 64 

cast, 56 

cells, 56 

cleaning pipettes, 62 

coagulation of, 68 

counters, 61 

counting fluids, 61 

development of cells, 53 

differential counts, 66 

diseases of, 58 

dramas, 55, 57 

forceps for, 65 

fowl's, 67 

hemin tests, 67 

hemoglobin, 59 

in exudates, 97 

in feces, 149 

in monoxide poisoning, 68 

in sputum, 32 

in stomach contents, 74 

in urine, 113 

leukocytes in, 57 

macroscopical examination of, 59 

microscopical examination of, 59 

obtaining specimen of, 58 

occult, 149 

parasites in, 67 

pipettes for, 61 

plates in, 54 

red cells of, 56 

red counting, 60 

spreading of, 63 

staining of, 65 

stickers for, 57 



Blood — cont'd. 

white cells of, 57 
counting, 62 

^Yidal tests of, 106 
Borax in milk, 135 
Boric acid in milk, 135 
Boston's test for albumin, 110 
Bothriocephalus latus in water, 141 
Bowel bacilli in water, 145 
Bracers, composition of, 87 
Brain in autopsy technic, 162 
Bronchitis, laboratory aids in, 184 
Burns, treatment of, 179 
Butyric Acid Test, 99 



Calcium carbonate, 120 

chlorid, 188 

as a reagent, 85 

oxalate, 120 

crystals in urine, 120 
oxaluria dolorosa, 120, 122 
Canada balsam, properties of, 188 
Cantharidin, detection of, 90 
Carbol-fuchsin, 31 

gentian violet, 31 

thionin, 81 

xylol, 84, 188 
Carbolic acid, 188 

detection of, 90 
Caries, 99 
Casts in urine, 117 
Celloidin, 188 

imbedding in, 82 
Centigrade equivalents, 193 
Centrifuge, 14 

choice, 14 

collecting sediment, 116 

use in exudates, 96 
Cerebro-spinal fever, laboratory aids 
in, 183 

fluid, 97 

obtaining, 94 
examination of, 97 
Charcoal, properties of, 188 
Chest, opening of, 164 
Chloral, detection of, 90 
Chlorides in urine, 115 
Chlorin estimation in water, 143 
Chloroform, properties of, 188 
Chronic rheumatism, 183 
Clap threads in urine, 117 
Cocam, 87 

detection of, 90 
Colon bacillus, 48 

in water, 145 
Commercial waters, 139 
Comparative chlorin, 143 
Copper, detection of, 91 



INDEX. 



203 



Cord; removal of, 168 

Cough syrups, composition of, 87 

Cover glasses, 16. 20 

sterilization of, 42 
Cream, proportion in milk, 134 
Crystals in urine. 120 
Culture media, 23, 27 
Curettings, 84 
Curschmann's spirals, 30 

in sputum, 33 
Cvlindroids in urine, 118 



D 



Dead animals in water, 141 
Decinormal sodium hydrate solution, 

75 
Degeneration of red corpuscles, 56 
Dengue, laboratory aids in, 183 
Dental caries. 99 
Deodorizers, 24, 194 
Detection of poisons, 85 

abortifacients, 87 

acetanilid, 88, 186 

acids, 88 

alcohol, 88, 187 

alkalies, 88 

ammonia, 88, 187 

antipyrin, 188 

apparatus for, 85 

arsenic, 89, 188 

atropin, 90, 188 

bad whisky, 87 

bitters, 87 

bracers, 87 

cantharidin, 90 

carbolic acid, 90, 186 

chloral hydrate, 90, 188 

cocain, 87, 90, 189 

cough syrups, 87 

drug cures, 87 

ergot, 91 

formalin, 91, 189 

headache powders, 87 

hydrocyanic acid, 91. 181, 187 

hyoscin, 90 

knock-out drops, 87 

lead. 91. 140. 190 

liniments, S7 

lye, 88 

matches, 88 

mercury salts, 91, 190 

morphin, 92, 190 

opium, 92, 190 

paris green, 88 

phenol. 90 

phosphorus, 92, 190 

rat poisons, 88 

sex stimulants, 88 

silver nitrate, 92, 191 



Detection — cont'd. 

skin beautifiers, 88 

sources of poisons, 87 

strvchnin, 92, 191 

sulphonal, 92, 191 

trional, 92 

wood alcohol, 187 
Diabetes mellitus, 184 

insipidus, 184 

urine, 123 
Diacetic acid, tests for, 116 
Diazo test for typhoid, 100 
Dimethylamidoazobenzol, 189 

in stomach analysis, 75 
Dimethylaminobenzaldehyde, 115 
Diphtheria, 183 

bacillus of, 44 
Diplococcus intracellulars meningi- 
tiditis, 97 

gonorrhea, 48 

pneumonia, 33 
Diseases, laboratory aids in, 182 

of alimentary tract of infant, 185 

of arteries, 185 

of blood, 184 

of ear, 184 

of eye, 185 

of heart, 185 

of intestine, 184 

of kidney, 184 

of larynx, 184 

of lung, 184 

of pericardium, 185 

of pharynx, 184 

of pleura, 184 

of spleen, 185 

of thymus, 185 

of thyroid, 185 
Disinfectants, definition of, 177 
Distilled water as a reagent, 189 
Drop method of estimating percent- 
age solutions, 192 
Drug cures, composition of, 87 
Duodenum, examination of, 167 

E 

Ecchinococcus in sputum, 33 
Ehrlich's diazo reaction, 100 
Elastic tissue in sputum, 29 
Endocarditis, malignant, precautions, 

179 
Eosin, 68, 84 
Eosinophilia, 58 
Ergot, detection of, 91 
Erysipelas, laboratory aids in, 183 
Erythrocytes, 56 

artifacts of, 64 

classification of, 56 

counting of, 60 

development of, 53 



204 



INDEX. 



Erythrocytes — cont'd, 

in exudates, 97 

in feces, 149, 154 

in sputum, 32 

in stomach contents, 73 

in urine, 113 
Essence of tissue diagnosis, 78 

apparatus for, 78, 82 

celloidin, 82 

curettings, 84 

freezer, 80 

imbedding, 83 

microtome, 79 

knife, 80 ' 

paraffin, 84 

sectioning, 80, 83 

staining, 80 
Ether, properties of, 189 
Ethyl chlorid, properties of, 189 
Ewald test breakfast, 71 
Explosive mixtures, 180 
Exudates, 94 

abdominal, 94 

apparatus for examining, 94 

lumbar, 94 

preparation of, 96 

thoracic, 94 



Fahrenheit equivalents, 193 
Family sterility, 97 
Fat estimation in milk, 135 
Fecal analysis, 148 

ameba coli, 150 

apparatus, 148 

ascaris lumbricoides, 150 

bacteria, 149 

blood, 149, 154 

color, 149 

consistency, 149 

hookworm, 152 

in infant, 154 

obtaining the specimen, 148 

occult blood, 149 

odor, 149 

ova, 153 

pinworms, 150 

tapeworms, 151 

value and limitations, 155 
Female genitalia, examination of, 168 
Fermentation test for glucose, 113 
Ferric chlorid solution, 189 

use as a reagent, 71 
Fluorescence test in water, 146 
Foam test for bilirubin, 113 
Food poisons, 183 
Foreign matter, 118 

in sputum, 30 

in urine, 118, 121 



Formalin, formaldehyde, 189 
detection in milk, 134 
detection in quantity, 91 
for hardening tissues, 80 

Formulas, milk, 136 

Freezer for tissues, 80 

Frozen sections, 79 



G 



Gall stones in feces, 155 
Gas, substitutes for, 17 
Gastric analysis, 71 

apparatus for, 71 

blood, 73 

chemical, 75 

color, 73 

interpretation of, 76 

macroscopic examination, 73 

microscopic examination, 74 

physical characteristics, 74 

preparation for, 71 

sources of error in, 77 

value and limitations of, 77 
General considerations, 13 

information, 186 
Gentian violet, 27 
Germicides, 177 
Germ growth, 38 

incubation as an aid, 41 

influences inhibiting, 38 

sterilization of, 41 
Germs, 37 

actinomyces, 49 

apparatus used in searching for, 37 

bacillus coli communis, 48 
diphtheria?, 44 
Koch -Weeks, 47 
tetani, 45 
typhosus, 48 

culture media for, 37 

difficulties in finding, 50 

gonococcus, 48 

incubation of, 41 

inoculations of, 40 

isolation of pure cultures, 40 

molds, 49 

staphylococci, 48 

streptococcus pyogenes, 47 
Glassware, 14 

cleaning of, 14 

sterilization of, 41 
Glucose in the urine, 112 
Glycerin, properties of, 189 
Gonococcus, searching for, 48 
Gonorrhea, laboratory aids in, 183 
Gout, laboratory aids in. 184 
Gunzburg's reagent in stomach analy- 
sis, 75 

properties of, 189 



INDEX. 



205 



Haines' sugar test, 112 
Hayem's fluid, 189 

for counting- erythrocytes, 59 
Hay fever, laboratory aids in, 184 
Headache powders, composition of, 87 
Heart, examination of ; 165 

failure cells in sputum, 33 
Heat and nitric acid test, 110 
Hemalum, use of, 84 
Hematuria, 119 
Hemoglobin, 59 

estimation of, 59 

in urine, 119 
Hermann-Perutz test, 199 
Hookworm, 152 

Hydrochloric acid (see Acids, Poi- 
sons, etc.) 
Hydrocyanic acid, cautions when in- 
haling, 181 

detection of, 91 

properties of, 187 
Hydrogen dioxid, 190 
Hydrophobia, laboratory aids in, 183 
Hyoscin, detection of, 90 

I 

Imbedding of tissues, 83 
Indican, significance of, 123, 195 

detection of, 196 
Indications for laboratory aids, 182 
Indicators, 15 

in gastric analysis, 75 

in urinalysis, 109 
Indolacetic acid, 197 
Infant feeding, 136 
Infant's stools, 154 
Inflammable chemicals, 181 
Influenza bacillus, 50 
Influenza, laboratory aids in, 183 
Ink methods to find the treponema 

pallidum, 171 
Inoculations, 38 

stabs and streaks, 38 
Internal myiasis, 182 
Internal psorospermiasis, 182 
Intestinal obstruction, laboratory aids 

in, 184 
Intestines, examination of, 167 
Iodin test for malingering, 124 
Iodoform as a deodorizer, 24 

properties of, 190 
Iodophilia, 58, 68 

J 
Jaundice, laboratory aids in, 184 

K 
Karyokinesis in blood cells, 56 
Karyorrhexis in blood cells, 56 



Kidneys, examination of, 166 
Klebs-Loffler bacillus, 45 
Knife for microtome, 80 
Knock-out drops, 87 



Laboratory aids in disease, 182 

arrangement of the physician's, 23 

bacteriological, 21 

books, 24 

equipment, 13 

experts, 25 

for milk analysis, 129 

prophylaxis, 177 

tables, 21, 23 
Lactic acid, test for, 75 
Lead acetate, 91 

in water, 140 
Leucorrhea, 98, 99 
Leukemia, 58 
Leukocytes, 57 

counting of, 62 

in exudates, 97 

in sputum, 32 

in urine, 117 
Leukocytosis, 58 
Leukopenia, 58 
Liniments, composition of, 87 
Litmus test in water, 146 
Liver, examination of, 167 
Lobar pneumonia, laboratory aids in, 

183 
Loffler's methylene blue, 45 : 48 
Lumbar puncture, 94 
Lungs, examination of, 166 
Lye, composition of, 88 
Lymphatic leukemia, 58 
Lymphocytosis, 97 

M 

Macroblast, 56 

Malaria, laboratory aids in, 182 

parasite of, 67 
Male genitalia, examination of, 167 
Marsh apparatus, 85 
Marx's fluid, 68 
Matches, composition of, 88 
Measles, laboratory aids in, 182 
Megaloblast, 56 
Megalocyte, 56 

Meninges, examination of, 161 
Mercurialism, 99 
Mercury salts, detection of, 91 
Methylene blue, use of, 44 
Microblast, 56 
Microcyte. 56 
Microscope, 14 

choice of instrument, 14 



206 



INDEX. 



Microscope — cont'd. 

condenser, 18 

focusing, 20 

illumination, 18 

iris diaphragm, 18 

lenses, 20 

mirror, 18 

objective, 20 

ocular, 19 
Microscopic hysteria, 20 

technic, 18 
Microtome, 79 

knife, 80 
Milk, 125 

advantages of modified, 128 

amount daily, 132 

analysis of, 133 

apparatus for analysis of, 125 

borax in,, 135 

color of, 133 

cream, 134 

fat estimation of, 135 

formaldehyde in, 134 

formulas, 136 

maternal, 137 

microscopy of, 134 

modifications of, 136 

odor of, 134 

prescription for, 129 

proprietary substitutes, 127 

reaction of, 134 

sample of, 131 

sodium bicarbonate in, 135 

specific gravity of, 134 

tampering with, 131 
Morphin, detection of, 92 
Mucus, 27 

in gastric contents, 74 

in sputum, 27 

in stools, 154 

in urine, 108 

versus pus, 27 
Mumps, laboratory aids in, 183 
Muscular rheumatism, laboratory aids 

in, 184 
Myelogenous leukemia, 58 

N 

Necropsy (see Autopsy) 
Negri bodies of hydrophobia, 183 
Neoplasms, laboratory aids in, 185 
Nessler's reagent, 90 

composition of, 190 
Neutral red in water analysis, 146 
New born, autopsy of, 169 
Normoblast, 56 



Obesity, laboratory aids in, 184 
Occult blood, test 'for, 149 



Oil of spearmint, 190 

Opium, detection of, 92 

Oppler-Boas bacillus, 75 

Order of procedure in autopsy, 159 

Ova of worms, 153 

Oxalates in urine, 121 

Oxaluria dolorosa, 121 



Pancreas, examination of, 167 
Pancreatic test, 114 
Paradoxical stain, 57 
Paraffin imbedding, 84 
Parasites, 151 

in blood, 67 

in feces, 151 

in urine, 121 

in water, 141 
Parasitic skin diseases, laboratory 

aids in, 185 
Paris green, composition of, 88 
Peritonitis, laboratory aids in, 184 
Phenol, detection of, 90 
Phenolphthalein, 15 

in stomach analysis, 75 

in urinalysis, 109 
Phosphates. 110 

crystals in urine, 121 
Phosphorus, detection of, 92 
Pinworms in feces, 150 
Pipettes, 16 

for blood counting. 53 
Plasmodium in the blood, 67 
Plate cultures of bacteria, 41 
Poikilocyte, 56 
Poisons, 85 

abortifacients, 87 

acetanilid, 88, 186 

acids, 88 

alkalies, 88 

alcohol, 88, 187 

ammonia, 88, 187 

antipyrin, 188 

apparatus for detection of, 85 

arsenic, 89, 188 

atropin, 90, 188 

bad whisky, 87 

bitters. 87 

bracers, 87 

cantharidin. 90 

carbolic acid. 90, 186 

chloral hydrate, 90, 188 

eocain, 87, 90, 189 

cough syrups, 87 

drug cures, 87 

ergot, 91 

formalin, 91, 189 

headache powders, 87 

hvdrocvanic acid, 91, 181, 187 



INDEX. 



207 



Poisons — con fd> 
hyosein, 90 

knock-out drops, 87 

lead, 91, 140, 190 

liniments, 87 

lye, 88 

matches, 88 

mercury salts, 91, 190 

morphin, 92, 190 

opium, 92, 190 

paris green, 88 

phenol, 90 

phosphorus, 92, 190 

rat poisons, 88 

sex stimulants, 88 

silver nitrate, 92, 191 

skin beautifiers, 88 

sources of, 87 

strychnin, 92, 191 

sulphonal, 92, 191 

trional, 92 

wood alcohol, 187 
Polychromatophilia, 57 
Post-mortem technic, 156 

bacteriological, 157 

brain, 162 

chest, 164 

cord, 168 

duodenum, 167 

equipment, 156 

female genitalia, 168 

heart, 165 

in new born, 169 

inspection, palpation, and percus- 
sion, 160 

intestines, 167 

kidneys, 166 

liver, 167 

lungs, 166 

male genitalia, 167 

meninges, 161 

microscopic morbid anatomy, 157' 

ordei of procedure, 159 

pancreas, 167 

precautions in, 156 

prepartions foi% 157 

private posi-m )rtem, 156 

prophylaxis Lr 179 

special sense oijans, 163 

spleen, 166, 

stomach, 167 

trunk, i o3 

uncover r e the Oram, 16 v 

weights aim r. .sures, 158 
Potassium oicnr mate, properties of, 
191 

chromate, pr e ties of ; 191 
as an ind T ' or 143 

hydrate, pr- t;?5 of, 191 
Pregnancy, lal » yry aids in, 185 



Prescriptions for modified milk, 
Proprietary infant foods, 127 
Proteids in milk, 130 

in urine, 109 
Ptyalism, 99 

Puncture fluids (see Exudates) 
Pus in exudates, 97 

in sputum, 27 

in urine, 117 

versus mucus, 27 



R 



Rape, proof of, 98 
Rat poisons, composition of, 88 
Reaction of gastric contents, 75 
of milk, 134 
of saliva, 99 
of urine, .109 
of water, 140 
Reagents, properties of, 186 
acacia, 186 
acid, acetic, 186 

chromic, 186 

hydrochloric, 187 

nitric, 187 

picric, 187 

sulphanilic, 187 

sulphuric, 187 
alcohol, ethyl, 187 

fusel, 187" 
ammonium hydrate, 187 
azolitmin, 188 
benzidin, 188' 
calcium chlorid, 188 
Canada balsam, 188 
carbol-xylol, 188 
celloidin, 188 
charcoal, 188 
chloroform, 188 
dimethylamidoazobenzoh 189 
distilled water, 189 
ether, 189 
ethyl chlorid, 189 
ferric chlorid, 189 
formalin, formaldehyde,, 189 
glycerin, 189 
Gunzburg's reagent, 189 
Haines' solution, 189 
Hayem's solution, 189 
hydrogen dioxid, 190 
iodoform, 190 
Marx's fluid, 68 
Nessler's reagent, 190 
oil of spearmint, 190 
phenolphthalein, 190 
potassium bichromate, 191 

chromate, 191 

hydrate, 191 
1 silver nitrate, 191 



129 



208 



INDEX. 



Reagents — cont'd. 

sodium chlorid, 191 
citrate, 191 
nitrite, 191 

stannous chlorid, 191 

sweet oil, 19 1 

turmeric, 191 

urotropin, 191 

zinc, 191 
Red blood corpuscles, 56 

artifacts in ; 64 

classification of, 56 

counting of, 60 

development of, 53 

in exudates, 97 

in feces, 149, 154 

in sputum, 32 

in stomach contents, 73 

in urine, 113 
Rheumatic fever, laboratory aids in, 

183 
Rickets, laboratory aids in 2 184 
Rubella, laboratory aids in, 183 
Russo typhoid test, 102 
Rust, removal of, 194 



Saccharimeter, 113 

Sapremia, laboratory aids in, 183 

Sarcines, 75 

significance of, 76 
Scarlet fever, laboratory aids in, 182 
Searching for germs (see Germs) 
Sectioning, 80, 83 
Septicemia, laboratory aids in, 183 
Serum proteins in urine, 109 
Sewage, 141 

Sex stimulants, composition of, 88 
Silver nitrate, detection of, 92 
Skin beautifiers, composition of, 88 
Slides, cleaning of, 14 
Smallpox, laboratory aids in, 184 
Smegma bacillus, differential stain, 

123 
Sodium chlorid, properties of, 191 

citrate, properties of, 191 
as a reagent, 96 

hydrate, solution of, 75 
Special sense organs, examination of, 

163 
Specific gravity of milk, 134 

of mine, 107 
Spermatozoa, identification of, 97 
Spermatorrhea, diagnosis of, 98 
Spirochete, 173 
Spleen, examination of, 166 
Spoon urinalysis, 194 
Sputum, 27 

apparatus for examination of, 27 



Sputum — cont'd. 

color of, 29 

consistence of, 27 

difficulties, 33 

elastic tissue in, 29 

leukocytes in, 32 

odor of, 29 

quantity of, 27 

red blood cells in, 32 

spreading of, 31 

staining of, 31 

tubercle bacillus in, 32 
Squibb's urea apparatus, 115 
Stab method of inoculation, 38 
Stains, 15 

carbol-fuchsin, 31 
gentian violet, 31 
thionin, 81 

eosin, 68, 84, 189 

Gram's, 84 

hemalum, 84, 190 

methylene blue, 48, 190 

neutral red, 190 

on fingers, 194 

simple, 43 

thionin, 81, 191 

Wright's, 65, 192 
Stannous chlorid, properties of, 191 
Staphylococcus, 48 
Starch grains in gastric analysis, 74 

in malingering test, 124 
Sterilization, 41 

of cover glasses, 42 

of glassware, 41 

of hands, 42 

of media, 41 

of metalware, 41 

of tables, 42 

of towels, 42 
Sterilizers, 22, 41 
Stomach analysis, 71 

apparatus for, 71 

blood, 73 

chemical, 75 

color, 73 

interpretation of, 76 

macroscopic examination, 73 

microscopic examination, 74 

physical characteristics, 74 

preparation for, 71 

sources of error in, 77 

value and limitations of, 77 
Stomach tube, use of, 72 

washing, technic of, 71 
Stomatitis, laboratory aids in, 184 
Stool tests, 148 

ameba coli, 150 

apparatus for, 148 

ascaris lumbrieoides, 150 

bacteria, 149 



INDEX. 



209 



Stool tests — cont'd. 

blood, 149, 154 

color, 140 

consistency, 149 

hookworm, 152 

infant's, 154 

Koch's bacillus, 150 

mucus, 154 

obtaining the specimen, 148 

occult blood, 149 

odor, 140 

ova, 153 

pinworms, 150 

tapeworms, 151 

value and limitations of, 155 
Streak method of inoculation, 38 
Streptococcus pyogenes, 47 
Strychnin, detection of, 02 
Sugar of milk, 130 

in urine, 113 
Sulphonal, detection of, 02 
Sulphosalicylic acid, 108 
Sweet oil as a reagent, 00 

properties of, 101 
Syphilis, laboratory aids in, 183 

butvric acid test for, 00 

Perutz test for, 100 



T 



Table microtome, 70 

Tallqvist hemoglobin chart, 50 

Tapeworms in feces, 151 

Tenia in feces, 151 

Test meals in gastric analysis, 71 

Tetanus, laboratory aids in, 183 

Thrush fungus, 40 

Tissues, examination of, 78 

apparatus, 78, 82 

celloidin, 82 

curettings, 84 

freezer, 80 

imbedding, 83 

microtome, 79 
knife, 80 

paraffin, 84 

sectioning of, 80, 83 

staining of, 80 
Tonsils, diseases of, 184 
Treponema examinations, 171 

advantages of methods in, 175 

apparatus necessary, 171 

criticism of methods, 174 

disadvantages of methods, 174 

examination in dry, 173 
in wet, 173 

identification, 173 

obtaining the specimen, 171 

preparing the specimen, 171 

principle involved, 175 

sources of error, 174 



Treponema examinations — cont'd 
spirochete as confusing elements, 
173 
Trional, detection of, 02 
Triple phosphates in urine, 121 
Trunk, examination of, 163 
Tubercle bacillus, 32 
in feces, 150 
in sputum, 32 
in urine, 121, 123 
Tuberculosis, laboratory aids in, 183 
Turmeric as a reagent, 135 

properties of, 101 
Typhoid bacillus, 48 

identification in drinking water, 
141 
fever, laboratory aids in, 182 
tests, 100 

U 

Urate of ammonium, 120 
Urea in urine, 115 
Uric acid in urine, 115, 120 
Urinary sediments, 116 

blood cells, 110 

calcium carbonate, 121 

calcium oxalate, 120 

casts, 117 

classification of, 120 

collection of, 116 

cylindroids, 118 

epithelial cells, 120 

leukocytes, 117 

phosphates, 121 

pus cells, 117 

triple phosphates, 121 

urates, 120 

uric acid, 120 

spermatozoa, 08 
Urine, 107 

albumin in, 100 

ammoniacal fermentation of, 121 

amount of, 107 

bacteria in, 123 

betaoxybutyric acid in, 116 

bilirubin in, 113 

blood in, 113 

casts in, 117 

characteristics of normal, 107 

chlorides in, 115 

choice of tests, 110 

clap threads in, 117 

color of, 108 

crystals in, 120 

cylindroids in, 118 

diacetic acid in, 116 

epithelium in, 120 

filtration of, 110 

foreign matter in, 118, 121 

glucose in, 112 

hemoglobin in, 110 



210 



INDEX. 



Urine — cont'd. 

leukocytes in, 117 

odor of, 108 

oxalates in, 120, 200 

parasites in, 121 

phosphates in, 110, 121 

pus in, 117 

reaction of, 109 

specific gravity of, 107 

spermatozoa in, 98 

tampering with, 124 

transparency of, 108 

turbidity of, 108 

urobilinogen in, 114 

worms in, 121 
Urine of acid cystitis, 122 

alkaline cystitis, 122 

autointoxication, 123 

diabetes, 123 

foudroyant cystitis, 122 

interstitial nephritis, 123 

oxaluria dolorosa, 121 

parenchymatous nephritis, 122 

pyelitis, 122 

renal calculus, 122 

specific urethritis, 122 

uremia, 122 

vesicle calculus, 122 
Urobilinogen, in urine, 114 
Urotropin as a substitute for gas, 17 

properties of, 191 
Uterine diseases, laboratory aids in, 
185 



Vaccinia, laboratory aids in, 182 
Varicella, laboratory aids in, 182 
Verrucse necrogenicse, 179 



W 

Water analysis, 138 
algae, 141 



Water — cont'd. 

animal parasites, 141 

apparatus, 144, 146 

bacteria, 142 

bowel bacilli, 145 

colon bacillus, 145 

commercial waters, 139 

comparative chlorin, 143 

dead animals, 141 

fluorescence test, 146 

lead, 140 

limitations of, 145 

litmus test in, 146 

odor, 140 

poisons, 139 

reaction, 140 

scientific versus practical, 138 

sewage, 142 

sources of error in, 144 

typhoid bacillus, 141 

value of, 145 
Water, changing of, 139 

running, for laboratory, 18 
Weights and measures, 192 

in post-mortem work, 158 
Whooping-cough, laboratory aids in, 

183 
Widal reaction of blood, 103 
Worms in feces, 150 
Wright's blood stain, 65, 192 



Yeasts in gastric contents, 75 
Yellow fever, laboratory aids in, 183 



Zinc, in arsenic test, 85 
in hemoglobin test, 120 
properties of, 191 



J 11 it uio 



